US12115228B2 - Drug-conjugates, conjugation methods, and uses thereof - Google Patents

Drug-conjugates, conjugation methods, and uses thereof Download PDF

Info

Publication number
US12115228B2
US12115228B2 US17/189,135 US202117189135A US12115228B2 US 12115228 B2 US12115228 B2 US 12115228B2 US 202117189135 A US202117189135 A US 202117189135A US 12115228 B2 US12115228 B2 US 12115228B2
Authority
US
United States
Prior art keywords
conjugate
active agent
pab
group
antibody
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Active, expires
Application number
US17/189,135
Other versions
US20210187119A1 (en
Inventor
Zhenwei Miao
Yufeng Hong
Tong Zhu
Alexander Wilhelm Chucholowski
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Vivasor Inc
Concortis Biosystems Corp
Original Assignee
Concortis Biosystems Corp
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Concortis Biosystems Corp filed Critical Concortis Biosystems Corp
Priority to US17/189,135 priority Critical patent/US12115228B2/en
Publication of US20210187119A1 publication Critical patent/US20210187119A1/en
Assigned to VIVASOR, INC. reassignment VIVASOR, INC. ASSIGNMENT OF ASSIGNORS INTEREST (SEE DOCUMENT FOR DETAILS). Assignors: SORRENTO THERAPEUTICS, INC.
Application granted granted Critical
Publication of US12115228B2 publication Critical patent/US12115228B2/en
Active legal-status Critical Current
Adjusted expiration legal-status Critical

Links

Images

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/68—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an antibody, an immunoglobulin or a fragment thereof, e.g. an Fc-fragment
    • A61K47/6835—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an antibody, an immunoglobulin or a fragment thereof, e.g. an Fc-fragment the modifying agent being an antibody or an immunoglobulin bearing at least one antigen-binding site
    • A61K47/6851—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an antibody, an immunoglobulin or a fragment thereof, e.g. an Fc-fragment the modifying agent being an antibody or an immunoglobulin bearing at least one antigen-binding site the antibody targeting a determinant of a tumour cell
    • A61K47/6855—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an antibody, an immunoglobulin or a fragment thereof, e.g. an Fc-fragment the modifying agent being an antibody or an immunoglobulin bearing at least one antigen-binding site the antibody targeting a determinant of a tumour cell the tumour determinant being from breast cancer cell
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/04—Peptides having up to 20 amino acids in a fully defined sequence; Derivatives thereof
    • A61K38/07—Tetrapeptides
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/68—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an antibody, an immunoglobulin or a fragment thereof, e.g. an Fc-fragment
    • A61K47/6801—Drug-antibody or immunoglobulin conjugates defined by the pharmacologically or therapeutically active agent
    • A61K47/6803—Drugs conjugated to an antibody or immunoglobulin, e.g. cisplatin-antibody conjugates
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/68—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an antibody, an immunoglobulin or a fragment thereof, e.g. an Fc-fragment
    • A61K47/6801—Drug-antibody or immunoglobulin conjugates defined by the pharmacologically or therapeutically active agent
    • A61K47/6803—Drugs conjugated to an antibody or immunoglobulin, e.g. cisplatin-antibody conjugates
    • A61K47/68031—Drugs conjugated to an antibody or immunoglobulin, e.g. cisplatin-antibody conjugates the drug being an auristatin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/68—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an antibody, an immunoglobulin or a fragment thereof, e.g. an Fc-fragment
    • A61K47/6801—Drug-antibody or immunoglobulin conjugates defined by the pharmacologically or therapeutically active agent
    • A61K47/6803—Drugs conjugated to an antibody or immunoglobulin, e.g. cisplatin-antibody conjugates
    • A61K47/6811—Drugs conjugated to an antibody or immunoglobulin, e.g. cisplatin-antibody conjugates the drug being a protein or peptide, e.g. transferrin or bleomycin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/68—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an antibody, an immunoglobulin or a fragment thereof, e.g. an Fc-fragment
    • A61K47/6835—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an antibody, an immunoglobulin or a fragment thereof, e.g. an Fc-fragment the modifying agent being an antibody or an immunoglobulin bearing at least one antigen-binding site
    • A61K47/6849—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an antibody, an immunoglobulin or a fragment thereof, e.g. an Fc-fragment the modifying agent being an antibody or an immunoglobulin bearing at least one antigen-binding site the antibody targeting a receptor, a cell surface antigen or a cell surface determinant
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/68—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an antibody, an immunoglobulin or a fragment thereof, e.g. an Fc-fragment
    • A61K47/6889—Conjugates wherein the antibody being the modifying agent and wherein the linker, binder or spacer confers particular properties to the conjugates, e.g. peptidic enzyme-labile linkers or acid-labile linkers, providing for an acid-labile immuno conjugate wherein the drug may be released from its antibody conjugated part in an acidic, e.g. tumoural or environment
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P3/00—Drugs for disorders of the metabolism
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P3/00—Drugs for disorders of the metabolism
    • A61P3/08—Drugs for disorders of the metabolism for glucose homeostasis
    • A61P3/10—Drugs for disorders of the metabolism for glucose homeostasis for hyperglycaemia, e.g. antidiabetics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders
    • A61P37/02—Immunomodulators
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders
    • A61P37/02—Immunomodulators
    • A61P37/04—Immunostimulants
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D207/00—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom
    • C07D207/02—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom
    • C07D207/04—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having no double bonds between ring members or between ring members and non-ring members
    • C07D207/08—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having no double bonds between ring members or between ring members and non-ring members with hydrocarbon radicals, substituted by hetero atoms, attached to ring carbon atoms
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D207/00—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom
    • C07D207/02—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom
    • C07D207/04—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having no double bonds between ring members or between ring members and non-ring members
    • C07D207/08—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having no double bonds between ring members or between ring members and non-ring members with hydrocarbon radicals, substituted by hetero atoms, attached to ring carbon atoms
    • C07D207/09—Radicals substituted by nitrogen atoms, not forming part of a nitro radical
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D277/00—Heterocyclic compounds containing 1,3-thiazole or hydrogenated 1,3-thiazole rings
    • C07D277/02—Heterocyclic compounds containing 1,3-thiazole or hydrogenated 1,3-thiazole rings not condensed with other rings
    • C07D277/20—Heterocyclic compounds containing 1,3-thiazole or hydrogenated 1,3-thiazole rings not condensed with other rings having two or three double bonds between ring members or between ring members and non-ring members
    • C07D277/587—Heterocyclic compounds containing 1,3-thiazole or hydrogenated 1,3-thiazole rings not condensed with other rings having two or three double bonds between ring members or between ring members and non-ring members with aliphatic hydrocarbon radicals substituted by carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to ring carbon atoms, said aliphatic radicals being substituted in the alpha-position to the ring by a hetero atom, e.g. with m >= 0, Z being a singly or a doubly bound hetero atom
    • C07D277/593—Z being doubly bound oxygen or doubly bound nitrogen, which nitrogen is part of a possibly substituted oximino radical
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
    • C07D401/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings
    • C07D401/12—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings linked by a chain containing hetero atoms as chain links
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D403/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00
    • C07D403/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing two hetero rings
    • C07D403/12—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing two hetero rings linked by a chain containing hetero atoms as chain links
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07F—ACYCLIC, CARBOCYCLIC OR HETEROCYCLIC COMPOUNDS CONTAINING ELEMENTS OTHER THAN CARBON, HYDROGEN, HALOGEN, OXYGEN, NITROGEN, SULFUR, SELENIUM OR TELLURIUM
    • C07F5/00—Compounds containing elements of Groups 3 or 13 of the Periodic Table
    • C07F5/02—Boron compounds
    • C07F5/025—Boronic and borinic acid compounds
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
    • C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
    • C07K5/06—Dipeptides
    • C07K5/06008—Dipeptides with the first amino acid being neutral
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
    • C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
    • C07K5/06—Dipeptides
    • C07K5/06008—Dipeptides with the first amino acid being neutral
    • C07K5/06017—Dipeptides with the first amino acid being neutral and aliphatic
    • C07K5/06034—Dipeptides with the first amino acid being neutral and aliphatic the side chain containing 2 to 4 carbon atoms
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
    • C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
    • C07K5/06—Dipeptides
    • C07K5/06008—Dipeptides with the first amino acid being neutral
    • C07K5/06017—Dipeptides with the first amino acid being neutral and aliphatic
    • C07K5/06034—Dipeptides with the first amino acid being neutral and aliphatic the side chain containing 2 to 4 carbon atoms
    • C07K5/06052—Val-amino acid
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
    • C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
    • C07K5/10—Tetrapeptides
    • C07K5/1002—Tetrapeptides with the first amino acid being neutral
    • C07K5/1005—Tetrapeptides with the first amino acid being neutral and aliphatic
    • C07K5/1008—Tetrapeptides with the first amino acid being neutral and aliphatic the side chain containing 0 or 1 carbon atoms, i.e. Gly, Ala
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
    • C07K7/04—Linear peptides containing only normal peptide links
    • C07K7/06—Linear peptides containing only normal peptide links having 5 to 11 amino acids

Definitions

  • an antibody or antibody fragment such as a single-chain variable fragment
  • ADC antibody-drug conjugate
  • Some embodiments provide active agent-conjugates, methods of preparing active agent-conjugates, and uses thereof.
  • the E-component includes a fragment selected from the group consisting of:
  • L 3 may be —(CH 2 )—; and L 4 may be —(CH 2 )—. In some embodiments, L 3 may be null; and L 4 may be null.
  • L 2 includes —(CH 2 ) n — where n may be 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. In some embodiments, L 2 includes —(CH 2 CH 2 O) n — where n may be 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. In some embodiments, L 2 includes Val-Cit-PAB, Val-Ala-PAB, Val-Lys(Ac)-PAB, Phe-Lys-PAB, Phe-Lys(Ac)-PAB, D-Val-Leu-Lys, Gly-Gly-Arg, Ala-Ala-Asn-PAB, Ala-PAB, or PAB.
  • L 2 includes peptide, oligosaccharide, —(CH 2 ) n —, —(CH 2 CH 2 O) n —, Val-Cit-PAB, Val-Ala-PAB, Val-Lys(Ac)-PAB, Phe-Lys-PAB, Phe-Lys(Ac)-PAB, D-Val-Leu-Lys, Gly-Gly-Arg, Ala-Ala-Asn-PAB, Ala-PAB, PAB, or combinations thereof.
  • L 2 includes a noncleavable unit.
  • the noncleavable unit includes —(CH 2 ) n — where n may be 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. In some embodiments, the noncleavable unit includes —(CH 2 CH 2 O) n — where n may be 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. In some embodiments, L 2 includes a cleavable unit. In some embodiments, the cleavable unit comprises a peptide.
  • the A component comprises a monoclonal antibody (mAB). In some embodiments, the A component comprises an antibody fragment, surrogate, or variant. In some embodiments, the A component comprises a protein ligand. In some embodiments, the A component comprises a protein scaffold. In some embodiments, the A component comprises a peptide. In some embodiments, the A component comprises a small molecule ligand. In some embodiments, the A component comprises at least one modified L-Alanine residue. In some embodiments, the A component comprises at least two modified L-Alanine residues. In some embodiments, at least one L 2 includes —(CH 2 ) n — where n may be 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.
  • At least one L 2 includes —(CH 2 CH 2 O) n — where n may be 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.
  • at least one L 2 includes Val-Cit-PAB, Val-Ala-PAB, Val-Lys(Ac)-PAB, Phe-Lys-PAB, Phe-Lys(Ac)-PAB, D-Val-Leu-Lys, Gly-Gly-Arg, Ala-Ala-Asn-PAB, Ala-PAB, or PAB.
  • At least one L 2 includes a peptide, an oligosaccharide, —(CH 2 ) n —, —(CH 2 CH 2 O) n —, Val-Cit-PAB, Val-Ala-PAB, Val-Lys(Ac)-PAB, Phe-Lys-PAB, Phe-Lys(Ac)-PAB, D-Val-Leu-Lys, Gly-Gly-Arg, Ala-Ala-Asn-PAB, Ala-PAB, PAB or combinations thereof.
  • FIG. 1 shows the HIC-HPLC chromatogram product of reacting drug and antibody at a drug/antibody ratio of 5.5:1.
  • FIG. 2 shows the HIC-HPLC chromatogram product of reacting drug and antibody at a drug/antibody ratio of 6:1.
  • FIG. 3 shows the HIC-HPLC chromatogram product of reacting drug and antibody at a drug/antibody ratio of 1:6.5.
  • FIG. 4 shows the HIC-HPLC chromatogram product of reacting drug and antibody at a drug/antibody ratio of 1:4
  • FIG. 5 shows HIC-HPLC chromatograms for each FPLC fraction, and, inset, an overlay of the peaks to demonstrate relative purity of each peak.
  • FIG. 6 shows reduced SDS-PAGE analysis for the FPLC fractions depicted in FIGS. 4 and 5 .
  • FIG. 7 shows viability of SK-BR-3 cells for seven antibody-drug conjugate samples.
  • FIG. 8 shows viability of HCC1954 cells for seven antibody-drug conjugate samples.
  • FIG. 9 shows reduced SDS-PAGE analysis for trastuzumab conjugated compound 40.
  • FIG. 10 shows viability of SK-BR-3 cells for four antibody-drug conjugate samples.
  • FIG. 11 shows viability of HCC1954 cells for four antibody-drug conjugate samples.
  • FIG. 12 shows viability of SK-BR-3 cells for four antibody-drug conjugate samples.
  • the active agent-conjugate is a drug-conjugate.
  • the drug-conjugate includes a targeting molecule.
  • the targeting molecule includes a monoclonal antibody (mAB).
  • the drug-conjugate includes a spacer or a multifunctional linker.
  • the spacer connects to the mAB by a sulfide bond.
  • the multifunctional linker connects to the mAB by a sulfide bond.
  • the spacer or multifunctional linker may be optionally connected to an auxiliary moiety.
  • the auxiliary moiety may be a second targeting molecule such as mAB and peptide.
  • the auxiliary moiety may be a hydrophilic polymer such as polyethylene glycol (PEG), and the like.
  • the spacer or multifunctional linker may include a 2- to 5-atom bridge. In some embodiments, the spacer or multifunctional linker may include a 4C (four carbon) bridge.
  • Conjugation methods to derivatize a polypeptide with a payload can be accomplished using a maleimido or vinyl moiety which can react with individual sulfhydryl group on an antibody via Michael addition reaction.
  • the free sulfhydryl group can be formed by reducing a disulfide bond in an antibody.
  • the structural integrity of an antibody can be compromised after opening disulfide bonds and attaching payloads to the exposed free thiols.
  • the compositions and methods provided herein provide conjugation through cysteine without decreased structural stability.
  • pharmaceutically acceptable salt refers to salts that retain the biological effectiveness and properties of a compound and, which are not biologically or otherwise undesirable for use in a pharmaceutical.
  • the compounds disclosed herein are capable of forming acid and/or base salts by virtue of the presence of amino and/or carboxyl groups or groups similar thereto.
  • Pharmaceutically acceptable acid addition salts can be formed with inorganic acids and organic acids. Inorganic acids from which salts can be derived include, for example, hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, and the like.
  • Organic acids from which salts can be derived include, for example, acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, salicylic acid, and the like.
  • Pharmaceutically acceptable base addition salts can be formed with inorganic and organic bases.
  • Inorganic bases from which salts can be derived include, for example, sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese, aluminum, and the like; particularly preferred are the ammonium, potassium, sodium, calcium and magnesium salts.
  • Organic bases from which salts can be derived include, for example, primary, secondary, and tertiary amines, substituted amines including naturally occurring substituted amines, cyclic amines, basic ion exchange resins, and the like, specifically such as isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, and ethanolamine. Many such salts are known in the art, as described in WO 87/05297, Johnston et al., published Sep. 11, 1987 (incorporated by reference herein in its entirety).
  • C a to C b or “C a-b ” in which “a” and “b” are integers refer to the number of carbon atoms in the specified group. That is, the group can contain from “a” to “b”, inclusive, carbon atoms.
  • a “C 1 to C 4 alkyl” or “C 1-4 alkyl” group refers to all alkyl groups having from 1 to 4 carbons, that is, CH 3 —, CH 3 CH 2 —, CH 3 CH 2 CH 2 —, (CH 3 ) 2 CH—, CH 3 CH 2 CH 2 CH 2 —, CH 3 CH 2 CH(CH 3 )— and (CH 3 ) 3 C—.
  • halogen or “halo,” as used herein, means any one of the radio-stable atoms of column 7 of the Periodic Table of the Elements, e.g., fluorine, chlorine, bromine, or iodine, with fluorine and chlorine being preferred.
  • alkyl refers to a straight or branched hydrocarbon chain that is fully saturated (i.e., contains no double or triple bonds).
  • the alkyl group may have 1 to 20 carbon atoms (whenever it appears herein, a numerical range such as “1 to 20” refers to each integer in the given range; e.g., “1 to 20 carbon atoms” means that the alkyl group may consist of 1 carbon atom, 2 carbon atoms, 3 carbon atoms, etc., up to and including 20 carbon atoms, although the present definition also covers the occurrence of the term “alkyl” where no numerical range is designated).
  • the alkyl group may also be a medium size alkyl having 1 to 9 carbon atoms.
  • the alkyl group could also be a lower alkyl having 1 to 4 carbon atoms.
  • the alkyl group may be designated as “C 1-4 alkyl” or similar designations.
  • C 1-4 alkyl indicates that there are one to four carbon atoms in the alkyl chain, i.e., the alkyl chain is selected from the group consisting of methyl, ethyl, propyl, iso-propyl, n-butyl, iso-butyl, sec-butyl, and t-butyl.
  • Typical alkyl groups include, but are in no way limited to, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, tertiary butyl, pentyl, hexyl, and the like.
  • alkoxy refers to the formula —OR wherein R is an alkyl as is defined above, such as “C 1-9 alkoxy”, including but not limited to methoxy, ethoxy, n-propoxy, 1-methylethoxy (isopropoxy), n-butoxy, iso-butoxy, sec-butoxy, and tert-butoxy, and the like.
  • alkylthio refers to the formula —SR wherein R is an alkyl as is defined above, such as “C 1-9 alkylthio” and the like, including but not limited to methylmercapto, ethylmercapto, n-propylmercapto, 1-methylethylmercapto (isopropylmercapto), n-butylmercapto, iso-butylmercapto, sec-butylmercapto, tert-butylmercapto, and the like.
  • alkenyl refers to a straight or branched hydrocarbon chain containing one or more double bonds.
  • the alkenyl group may have 2 to 20 carbon atoms, although the present definition also covers the occurrence of the term “alkenyl” where no numerical range is designated.
  • the alkenyl group may also be a medium size alkenyl having 2 to 9 carbon atoms.
  • the alkenyl group could also be a lower alkenyl having 2 to 4 carbon atoms.
  • the alkenyl group may be designated as “C 2-4 alkenyl” or similar designations.
  • C 2-4 alkenyl indicates that there are two to four carbon atoms in the alkenyl chain, i.e., the alkenyl chain is selected from the group consisting of ethenyl, propen-1-yl, propen-2-yl, propen-3-yl, buten-1-yl, buten-2-yl, buten-3-yl, buten-4-yl, 1-methyl-propen-1-yl, 2-methyl-propen-1-yl, 1-ethyl-ethen-1-yl, 2-methyl-propen-3-yl, buta-1,3-dienyl, buta-1,2,-dienyl, and buta-1,2-dien-4-yl.
  • Typical alkenyl groups include, but are in no way limited to, ethenyl, propenyl, butenyl, pentenyl, and hexenyl, and the like.
  • alkynyl refers to a straight or branched hydrocarbon chain containing one or more triple bonds.
  • the alkynyl group may have 2 to 20 carbon atoms, although the present definition also covers the occurrence of the term “alkynyl” where no numerical range is designated.
  • the alkynyl group may also be a medium size alkynyl having 2 to 9 carbon atoms.
  • the alkynyl group could also be a lower alkynyl having 2 to 4 carbon atoms.
  • the alkynyl group may be designated as “C 2-4 alkynyl” or similar designations.
  • C 2-4 alkynyl indicates that there are two to four carbon atoms in the alkynyl chain, i.e., the alkynyl chain is selected from the group consisting of ethynyl, propyn-1-yl, propyn-2-yl, butyn-1-yl, butyn-3-yl, butyn-4-yl, and 2-butynyl.
  • Typical alkynyl groups include, but are in no way limited to, ethynyl, propynyl, butynyl, pentynyl, and hexynyl, and the like.
  • aromatic refers to a ring or ring system having a conjugated pi electron system and includes both carbocyclic aromatic (e.g., phenyl) and heterocyclic aromatic groups (e.g., pyridine).
  • carbocyclic aromatic e.g., phenyl
  • heterocyclic aromatic groups e.g., pyridine
  • the term includes monocyclic or fused-ring polycyclic (i.e., rings which share adjacent pairs of atoms) groups provided that the entire ring system is aromatic.
  • aryloxy and arylthio refers to RO- and RS-, in which R is an aryl as is defined above, such as “C 6-10 aryloxy” or “C 6-10 arylthio” and the like, including but not limited to phenyloxy.
  • an “aralkyl” or “arylalkyl” is an aryl group connected, as a substituent, via an alkylene group, such as “C 7-14 aralkyl” and the like, including but not limited to benzyl, 2-phenylethyl, 3-phenylpropyl, and naphthylalkyl.
  • the alkylene group is a lower alkylene group (i.e., a C 1-4 alkylene group).
  • heteroaryl refers to an aromatic ring or ring system (i.e., two or more fused rings that share two adjacent atoms) that contain(s) one or more heteroatoms, that is, an element other than carbon, including but not limited to, nitrogen, oxygen and sulfur, in the ring backbone.
  • heteroaryl is a ring system, every ring in the system is aromatic.
  • the heteroaryl group may have 5-18 ring members (i.e., the number of atoms making up the ring backbone, including carbon atoms and heteroatoms), although the present definition also covers the occurrence of the term “heteroaryl” where no numerical range is designated.
  • the heteroaryl group has 5 to 10 ring members or 5 to 7 ring members.
  • the heteroaryl group may be designated as “5-7 membered heteroaryl,” “5-10 membered heteroaryl,” or similar designations.
  • heteroaryl rings include, but are not limited to, furyl, thienyl, phthalazinyl, pyrrolyl, oxazolyl, thiazolyl, imidazolyl, pyrazolyl, isoxazolyl, isothiazolyl, triazolyl, thiadiazolyl, pyridinyl, pyridazinyl, pyrimidinyl, pyrazinyl, triazinyl, quinolinyl, isoquinlinyl, benzimidazolyl, benzoxazolyl, benzothiazolyl, indolyl, isoindolyl, and benzothienyl.
  • heteroarylkyl or “heteroarylalkyl” is heteroaryl group connected, as a substituent, via an alkylene group. Examples include but are not limited to 2-thienylmethyl, 3-thienylmethyl, furylmethyl, thienylethyl, pyrrolylalkyl, pyridylalkyl, isoxazollylalkyl, and imidazolylalkyl.
  • the alkylene group is a lower alkylene group (i.e., a C 1-4 alkylene group).
  • carbocyclyl means a non-aromatic cyclic ring or ring system containing only carbon atoms in the ring system backbone. When the carbocyclyl is a ring system, two or more rings may be joined together in a fused, bridged or spiro-connectedfashion. Carbocyclyls may have any degree of saturation provided that at least one ring in a ring system is not aromatic. Thus, carbocyclyls include cycloalkyls, cycloalkcnyls, and cycloalkynyls.
  • the carbocyclyl group may have 3 to 20 carbon atoms, although the present definition also covers the occurrence of the term “carbocyclyl” where no numerical range is designated.
  • the carbocyclyl group may also be a medium size carbocyclyl having 3 to 10 carbon atoms.
  • the carbocyclyl group could also be a carbocyclyl having 3 to 6 carbon atoms.
  • the carbocyclyl group may be designated as “C 3-6 carbocyclyl” or similar designations.
  • carbocyclyl rings include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cyclohexenyl, 2,3-dihydro-indene, bicycle[2.2.2]octanyl, adamantyl, and Spiro[4.4]nonanyl
  • a “(carbocyclyl)alkyl” is a carbocyclyl group connected, as a substituent, via an alkylene group, such as “C 4-10 (carbocyclyl)alkyl” and the like, including but not limited to, cyclopropylmethyl, cyclobutylmethyl, cyclopropylethyl, cyclopropylbutyl, cyclobutylethyl, cyclopropylisopropyl, cyclopentylmethyl, cyclopentylethyl, cyclohexylmethyl, cyclohexylcthyl, cycloheptylmethyl, and the like.
  • the alkylene group is a lower alkylene group.
  • cycloalkyl means a fully saturated carbocyclyl ring or ring system. Examples include cyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl.
  • cycloalkenyl means a carbocyclyl ring or ring system having at least one double bond, wherein no ring in the ring system is aromatic.
  • An example is cyclohexenyl.
  • heterocyclyl means a non-aromatic cyclic ring or ring system containing at least one heteroatom in the ring backbone. Heterocyclyls may be joined together in a fused, bridged or spino-connected fashion. Heterocyclyls may have any degree of saturation provided that at least one ring in the ring system is not aromatic. The heteroatom(s) may be present in either a non-aromatic or aromatic ring in the ring system.
  • the heterocyclyl group may have 3 to 20 ring members (i.e., the number of atoms making up the ring backbone, including carbon atoms and heteroatoms), although the present definition also covers the occurrence of the term “heterocyclyl” where no numerical range is designated.
  • the heterocyclyl group may also be a medium size heterocyclyl having 3 to 10 ring members.
  • the heterocyclyl group could also be a heterocyclyl having 3 to 6 ring members.
  • the heterocyclyl group may be designated as “3-6 membered heterocyclyl” or similar designations.
  • the heteroatom(s) are selected from one up to three of O (oxygen), N (nitrogen) or S (sulfur), and in preferred five membered monocyclic heterocyclyls, the heteroatom(s) are selected from one or two heteroatoms selected from O (oxygen), N (nitrogen), or S (sulfur).
  • heterocyclyl rings include, but are not limited to, azepinyl, acridinyl, carbazolyl, cinnolinyl, dioxolanyl, imidazolinyl, imidazolidinyl, morpholinyl, oxiranyl, oxepanyl, thiepanyl, piperidinyl, piperazinyl, dioxopiperazinyl, pyrrolidinyl, pyrrolidonyl, pyrrolidionyl, 4-piperidonyl, pyrazolinyl, pyrazolidinyl, 1,3-dioxinyl, 1,3-dioxanyl, 1,4-dioxinyl, 1,4-dioxanyl, 1,3-oxathianyl, 1,4-oxathiinyl, 1,4-oxathianyl, 2H-1,2-oxazinyl, trioxanyl, hexahydr
  • a “(heterocyclyl)alkyl” is a heterocyclyl group connected, as a substituent, via an alkylene group. Examples include, but are not limited to, imidazolinylmethyl and indolinylethyl.
  • acyl refers to —C( ⁇ O)R, wherein R is hydrogen, C 1-6 alkyl, C 2-6 alkenyl, C 2-6 alkynyl, C 3-7 carbocyclyl, C 6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
  • R is hydrogen, C 1-6 alkyl, C 2-6 alkenyl, C 2-6 alkynyl, C 3-7 carbocyclyl, C 6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
  • Non-limiting examples include formyl, acetyl, propanoyl, benzoyl, and acryl.
  • An “O-carboxy” group refers to a “—OC( ⁇ O)R” group in which R is selected from hydrogen, C 1-6 alkyl, C 2-6 alkenyl, C 2-6 alkynyl, C 3-7 carbocyclyl, C 6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
  • C-carboxy refers to a “—C( ⁇ O)OR” group in which R is selected from hydrogen, C 1-6 alkyl, C 2-6 alkenyl, C 2-6 alkynyl, C 3-7 carbocyclyl, C 6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
  • R is selected from hydrogen, C 1-6 alkyl, C 2-6 alkenyl, C 2-6 alkynyl, C 3-7 carbocyclyl, C 6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
  • a non-limiting example includes carboxyl (i.e., —C( ⁇ O)OH).
  • a “cyano” group refers to a “—CN” group.
  • a “cyanato” group refers to an “—OCN” group.
  • An “isocyanato” group refers to a “—NCO” group.
  • a “thiocyanato” group refers to a “—SCN” group.
  • An “isothiocyanato” group refers to an “—NCS” group.
  • a “sulfinyl” group refers to an “—S( ⁇ O)R” group in which R is selected from hydrogen, C 1-6 alkyl, C 2-6 alkenyl, C 2-6 alkynyl, C 3-7 carbocyclyl, C 6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
  • a “sulfonyl” group refers to an “—SO 2 R” group in which R is selected from hydrogen, C 1-6 alkyl, C 2-6 alkenyl, C 2-6 alkynyl, C 3-7 carbocyclyl, C 6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
  • S-sulfonamido refers to a “—SO 2 NR A R B ” group in which R A and R B are each independently selected from hydrogen, C 1-6 alkyl, C 2-6 alkenyl, C 2-6 alkynyl, C 3-7 carbocyclyl, C 6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
  • N-sulfonamido refers to a “—N(R A )SO 2 R B ” group in which R A and R b are each independently selected from hydrogen, C 1-6 alkyl, C 2-6 alkenyl, C 2-6 alkynyl, C 3-7 carbocyclyl, C 6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
  • An “O-carbamyl” group refers to a “—OC( ⁇ O)NR A R B ” group in which R A and R B are each independently selected from hydrogen, C 1-6 alkyl, C 2-6 alkenyl, C 2-6 alkynyl, C 3-7 carbocyclyl, C 6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
  • N-carbamyl refers to an “—N(R A )C( ⁇ O)OR B ” group in which R A and R B are each independently selected from hydrogen, C 1-6 alkyl, C 2-6 alkenyl, C 2-6 alkynyl, C 3-7 carbocyclyl, C 6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
  • An “O-thiocarbamyl” group refers to a “—OC( ⁇ S)NR A R B ” group in which R A and R B are each independently selected from hydrogen, C 1-6 alkyl, C 2-6 alkenyl, C 2-6 alkynyl, C 3-7 carbocyclyl, C 6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
  • a “urea” group refers to a “—N(R A )C( ⁇ O)NR A R B ” group in which R A and R B are each independently selected from hydrogen, C 1-6 alkyl, C 2-6 alkenyl, C 2-6 alkynyl, C 3-7 carbocyclyl, C 6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
  • N-thiocarbamyl refers to an “—N(R A )C( ⁇ S)OR B ” group in which R A and R B are each independently selected from hydrogen, C 1-6 alkyl, C 2-6 alkenyl, C 2-6 alkynyl, C 3-7 carbocyclyl, C 6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
  • a “C-amido” group refers to a “—C( ⁇ O)NR A R B ” group in which R A and R B are each independently selected from hydrogen, C 1-6 alkyl, C 2-6 alkenyl, C 2-6 alkynyl, C 3-7 carbocyclyl, C 6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
  • N-amido refers to a “—N(R A )C( ⁇ O)R B ” group in which R A and R B are each independently selected from hydrogen, C 1-6 alkyl, C 2-6 alkenyl, C 2-6 alkynyl, C 3-7 carbocyclyl, C 6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
  • amino group refers to a “—NR A R B ” group in which R A and R B are each independently selected from hydrogen, C 1-6 alkyl, C 2-6 alkenyl, C 2-6 alkynyl, C 3-7 carbocyclyl, C 6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
  • R A and R B are each independently selected from hydrogen, C 1-6 alkyl, C 2-6 alkenyl, C 2-6 alkynyl, C 3-7 carbocyclyl, C 6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
  • a non-limiting example includes free amino (i.e., —NH 2 ).
  • aminoalkyl refers to an amino group connected via an alkylene group.
  • alkoxyalkyl refers to an alkoxy group connected via an alkylene group, such as a “C 2-8 alkoxyalkyl” and the like.
  • a substituted group is derived from the unsubstituted parent group in which there has been an exchange of one or more hydrogen atoms for another atom or group.
  • substituted it is meant that the group is substituted with one or more substitutents independently selected from C 1 -C 6 alkyl, C 2 -C 6 alkenyl, C 2 -C 6 alkynyl, C 3 -C 7 carbocyclyl (optionally substituted with halo, C 1 -C 6 alkyl, C 1 -C 6 alkoxy, C 1 -C 6 haloalkyl, and C 1 -C 6 haloalkoxy), C 3 -C 7 -carbocyclyl-C 1 -C 6 -alkyl (optionally substituted with halo, C 1 -C 6 alkyl, C 1 -C 6 alkoxy, C 1 -C 6 haloalkyl
  • radical naming conventions can include either a mono-radical or a di-radical, depending on the context.
  • a substituent requires two points of attachment to the rest of the molecule, it is understood that the substituent is a di-radical.
  • a substituent identified as alkyl that requires two points of attachment includes di-radicals such as —CH 2 —, —CH 2 CH 2 —, —CH 2 CH(CH 3 )CH 2 —, and the like.
  • Other radical naming conventions clearly indicate that the radical is a di-radical such as “alkylene” or “alkenylene.”
  • a substituent is depicted as a di-radical (i.e., has two points of attachment to the rest of the molecule), it is to be understood that the substituent can be attached in any directional configuration unless otherwise indicated.
  • Subject as used herein, means a human or a non-human mammal, e.g., a dog, a cat, a mouse, a rat, a cow, a sheep, a pig, a goat, a non-human primate or a bird, e.g., a chicken, as well as any other vertebrate or invertebrate.
  • Some embodiments provide a method of conjugating of a targeting molecule through a spacer or a multifunctional linker.
  • the spacer or multifunctional linker may include a 2- to 5-atom bridge.
  • the method includes a single-step or sequential conjugation approach.
  • the drug-conjugates include a spacer or a multifunctional linker.
  • the spacer or multifunctional linker may include a noncleavable or cleavable unit such as peptides.
  • Some embodiments provide a method of treating a patient in need thereof comprising administering an active agent-conjugate as disclosed and described herein to said patient.
  • the patient may have cancer, immune diseases or diabetes.
  • Some embodiments provide a method of diagnosis or imaging comprising administering an active agent-conjugate as disclosed and described herein to an individual.
  • A may be a monoclonal antibody (mAB).
  • A may be an antibody fragment, surrogate, or variant.
  • A may be a protein ligand.
  • A may be a protein scaffold.
  • A may be a peptide
  • A may be a small molecule ligand.
  • B may be a hydrophilic polymer.
  • the hydrophilic polymer may polyethylene glycol (PEG), and the like.
  • B may be a biodegradable polymer.
  • the biodegradable polymer may be unstructured proteins polyamino acids, polypeptides polysaccharides and combinations thereof.
  • B may be a monoclonal antibody (mAB).
  • B may be an antibody fragment, surrogate, or variant.
  • B may be a protein ligand.
  • B may be a protein scaffold.
  • B may be a peptide
  • B may be RNA or DNA.
  • B may be a RNA or DNA fragment.
  • B may be a small molecule ligand.
  • D may be a biologically active compound.
  • D may be a drug
  • D may be a chemotherapy drug.
  • D may be a natural product.
  • D may be an immune modulator.
  • D may be a tubulin-binder
  • D may be a DNA-alkylating agent.
  • D may be an HSP90 inhibitor.
  • D may be a DNA topoisomerase inhibitor.
  • D may be an anti-epigenetic agent.
  • D may be an HDAC inhibitor.
  • D may be an anti-metabolism agent.
  • D may be a proteasome inhibitor.
  • D may be a peptide
  • D may be a peptidomimetic
  • D may be an siRNA.
  • D may be an antisense DNA.
  • D may be epothilone A, epothilone B, or paclitaxel.
  • L 2 may include a spacer or a multifunctional linker. In some embodiments, L 2 may include a spacer and a multifunctional linker. In some embodiments, L 2 may include a multifunctional linker. In some embodiments, each L 2 may be a linker, wherein the linker may be cleavable or non-cleavable under biological conditions. In some embodiments, the linker may be cleavable by an enzyme. In some embodiments, L 2 may include Linker.
  • L 1 includes a 2- to 5-carbon bridge and at least one sulfur atom. In some embodiments, L 1 includes a 2- to 5-carbon bridge and at least two sulfur atoms. In some embodiments, L 1 includes a 2- to 5-carbon bridge and a spacer. In some embodiments, L 1 includes a 2- to 5-carbon bridge, at least two sulfur atoms and a spacer. In some embodiments, L 1 may include one or more sulfurs. In some embodiments, the L 1 may include two or more sulfurs. In some embodiments, the L 1 may include exactly two sulfurs. In some embodiments, may include a 4-carbon bridge and/or a spacer. In some embodiments, L 1 include a 4-carbon bridge or a spacer.
  • L 1 may include a 4-carbon bridge and a spacer. In some embodiments, L 1 includes a 4-carbon bridge and at least two sulfur atoms. In some embodiments, the spacer connects to the mAB by a sulfide bond. In some embodiments, the spacer connects to the mAB through a thioether.
  • A comprises at least one modified L-Alanine residue. In some embodiments, A comprises at least two modified L-Alanine residues. In some embodiments, L 1 comprises at least one sulfur. In some embodiments, L 1 comprises at least two sulfurs. In some embodiments, A comprises at least one modified L-Alanine residue that is connected to at least one sulfur of L 1 . In some embodiments, at least one modified L-Alanine residue is from an L-Cysteine residue of a peptide before conjugation. In some embodiments, at least one sulfur of L 1 is from an L-Cysteine of a peptide before conjugation. In some embodiments, A and L 1 together comprise at least one thioether.
  • Linker may be a peptide
  • Linker may include an oligosaccharide.
  • Linker may include chitosan.
  • L 2 may include Linker and —(CH 2 ) n — where n is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.
  • L 2 may include Linker and —(CH 2 CH 2 O) n — where n is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.
  • Linker may include —(CH 2 ) n — where n is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.
  • Linker may include —(CH 2 CH 2 O) n — where n is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.
  • Linker may include Val-Cit-PAB, Val-Ala-PAB, Val-Lys(Ac)-PAB, Phe-Lys-PAB, Phe-Lys(Ac)-PAB, D-Val-Leu-Lys, Gly-Gly-Arg, Ala-Ala-Asn-PAB, Ala-PAB, PAB, or the like.
  • Linker may include any combination of peptide, oligosaccharide, —(CH 2 ) n —, —(CH 2 CH 2 O) n —, Val-Cit-PAB, Val-Ala-PAB, Val-Lys(Ac)-PAB, Phe-Lys-PAB, Phe-Lys(Ac)-PAB, D-Val-Leu-Lys, Gly-Gly-Arg, Ala-Ala-Asn-PAB, Ala-PAB, PAB, and the like.
  • the spacer may include a peptide.
  • the spacer may include an oligosaccharide.
  • the spacer may include chitosan.
  • the spacer may include —(CH 2 ) n — where n is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.
  • L 1 may include a component including a 4-carbon bridge and —(CH 2 ) n — where n is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.
  • the spacer may include —(CH 2 CH 2 O) n — where n is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.
  • L 1 may include a component including a 4-carbon bridge and —(CH 2 CH 2 O) n — where n is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.
  • the spacer may include Val-Cit-PAB, Val-Ala-PAB, Val-Lys(Ac)-PAB, Phe-Lys-PAB, Phe-Lys(Ac)-PAB, D-Val-Leu-Lys, Gly-Gly-Arg, Ala-Ala-Asn-PAB, Ala-PAB, PAB, or the like.
  • the spacer may be any combination of peptide, oligosaccharide, —(CH 2 ) n —, —(CH 2 CH 2 O) n —, Val-Cit-PAB, Val-Ala-PAB, Val-Lys(Ac)-PAB, Phe-Lys-PAB, Phe-Lys(Ac)-PAB, D-Val-Leu-Lys, Gly-Gly-Arg, Ala-Ala-Asn-PAB, Ala-PAB, PAB, and the like.
  • L 1 may include a 4-carbon bridge. In some embodiments, L 1 may include a 4-carbon bridge through a fused aromatic ring. In some embodiments, L 1 may include a spacer and component including a 4-carbon bridge through a fused aromatic ring. In some embodiments, L 1 may include, but is not limited
  • the agent-conjugate having the structure of Formula I has the structure of Formula Ia:
  • the S-linked (sulfur-linked) portion of A comprises a modified L-Alanine residue. In some embodiments, the S-linked (sulfur-linked) portion of A comprises a modified L-Alanine residue that is connected to the sulfur of L 1 . In some embodiments, the S-linked (sulfur-linked) portion of A comprises a modified L-Alanine residue wherein the modified L-Alanine component of A is from an L-Cysteine residue of a peptide before conjugation. In some embodiments, the sulfur of L 1 is from an L-Cysteine of a peptide before conjugation. In some embodiments, A and L 1 comprise a thioether. In some embodiments, the compound having the structure of Formula I a may be formed from a peptide that includes at least one L-Cysteine where the L-Cysteine provides a modified L-Alanine of A and a sulfur of
  • the agent-conjugate having the structure of Formula I has the structure of Formula Ib:
  • the E-component includes a fragment selected from the group consisting of:
  • the active agent may be selected from the group consisting of tubulin binders, DNA alkylators, DNA intercalator, enzyme inhibitors, immune modulators, peptides, and nucleotides.
  • At least one L 2 includes —(CH 2 ) n — where n is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. In some embodiments, at least one L 2 includes —(CH 2 CH 2 O) n — where n is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. In some embodiments, at least one L 2 includes Val-Cit-PAB, Val-Ala-PAB, Val-Lys(Ac)-PAB, Phe-Lys-PAB, Phe-Lys(Ac)-PAB, D-Val-Leu-Lys, Gly-Gly-Arg, Ala-Ala-Asn-PAB, Ala-PAB, or PAB.
  • At least one L 2 includes a peptide, an oligosaccharide, —(CH 2 ) n —, —(CH 2 CH 2 O) n —, Val-Cit-PAB, Val-Ala-PAB, Val-Lys(Ac)-PAB, Phe-Lys-PAB, Phe-Lys(Ac)-PAB, D-Val-Leu-Lys, Gly-Gly-Arg, Ala-Ala-Asn-PAB, Ala-PAB, or PAB.
  • at least one L 2 may comprise, consist of, or consist essentially of:
  • the agent-conjugate having the structure of Formula I has the structure of
  • the agent-conjugate having the structure of Formula I has the structure of
  • the agent-conjugate having the structure of Formula I has the structure of
  • the agent-conjugate having the structure of Formula I has the structure of
  • the agent-conjugate having the structure of Formula I has the structure of
  • the agent-conjugate having the structure of Formula I has the structure of
  • the agent-conjugate having the structure of Formula I has the structure of
  • the agent-conjugate having the structure of Formula I has the structure of
  • the agent-conjugate having the structure of Formula I has the structure of
  • the agent-conjugates may include one or more components selected from the group consisting of an amino acid, an amino acid residue, an amino acid analog, and a modified amino acid.
  • targeting moiety refers to a structure that binds or associates with a biological moiety or fragment thereof.
  • the targeting moiety may be an antibody. In some embodiments, the targeting moiety may be a monoclonal antibody (mAB). In some embodiments, the targeting moiety may be an antibody fragment, surrogate, or variant. In some embodiments, the targeting moiety may be a protein ligand. In some embodiments, the targeting moiety may be a protein scaffold. In some embodiments, the targeting moiety may be a peptide. In some embodiments, the targeting moiety may be RNA or DNA. In some embodiments, the targeting moiety may be a RNA or DNA fragment. In some embodiments, the targeting moiety may be a small molecule ligand.
  • mAB monoclonal antibody
  • the targeting moiety may be an antibody fragment, surrogate, or variant.
  • the targeting moiety may be a protein ligand. In some embodiments, the targeting moiety may be a protein scaffold. In some embodiments, the targeting moiety may be a peptide. In some embodiments, the targeting moiety may be RNA or DNA. In some
  • the targeting moiety may be an antibody fragment described in Janthur et al., “Drug Conjugates Such as Antibody Drug Conjugates (ADCs), Immunotoxins and Immunoliposomes Challenge Daily Clinical Practice,” Int. J. Mol. Sci. 2012, 13, 16020-16045, the disclosure of which is incorporated herein by reference in its entirety.
  • the targeting moiety may be an antibody fragment described in Trail, PA, “Antibody Drug Conjugates as Cancer Therapeutics,” Antibodies 2013, 2, 113-129, the disclosure of which is incorporated herein by reference in its entirety.
  • the targeting moiety may be HuM195-Ac-225, HuM195-Bi-213, Anyara (naptumomab estafenatox; ABR-217620), AS1409, Zevalin (ibritumomab tiuxetan), BIIB015, BT-062, Neuradiab, CDX-1307, CR011-vcMMAE, Trastuzumab-DM1 (R3502), Bexxar (tositumomab), IMGN242, IMGN388, 1MGN901, 131 I-labetuzumab, IMMU-102 ( 90 Y-epratuzumab), IMMU-107 ( 90 Y-clivatuzumab tetraxetan), MDX-1203, CAT-8015, EMD 273063 (hu14.18-1L2), Tucotuzumab celmoleukin (EMD 273066; huKS-
  • the targeting moiety may comprise, consist of, or consist essentially of the antibody portion of HuM195-Ac-225, HuM195-Bi-213, Anyara (naptumomab estafenatox; ABR-217620), AS1409, Zevalin (ibritumomab tiuxetan), BIIB015, BT-062, Neuradiab, CDX-1307, CR011-vcMMAE, Trastuzumab-DM1 (R3502), Bexxar (tositumomab), IMGN242, IMGN388, IMGN901, 131 I-labetuzumab, IMMU-102 ( 90 Y-epratuzumab), IMMU-107 ( 90 Y-clivatuzumab tetraxetan), MDX-1203, CAT-8015, EMD 273063 (hu14.18-1L2), Tucotuzumab celmoleuk
  • the targeting moiety may be Brentuximab vedotin, Trastuzumab emtansine, Inotuzumab ozogamicin, Lorvotuzumab mertansine, Glembatumumab vedotin, SAR3419, Moxetumomab pasudotox, Moxetumomab pasudotox, AGS-16M8F, AGS-16M8F, BIIB-015, BT-062, IMGN-388, or IMGN-388.
  • the targeting moiety may comprise, consist of, or consist essentially of the antibody portion of Brentuximab vedotin, Trastuzumab emtansine, Inotuzumab ozogamicin, Lorvotuzumab mertansine, Glembatumumab vedotin, SAR3419, Moxetumomab pasudotox, Moxetumomab pasudotox, AGS-16M8F, AGS-16M8F, BIIB-015, BT-062, IMGN-388, or IMGN-388.
  • the targeting moiety may comprise, consist of, or consist essentially of Brentuximab, Inotuzumab, Gemtuzumab, Milatuzumab, Trastuzumab, Glembatumomab, Lorvotuzumab, or Lab estuzumab.
  • peptide refers to a structure including one or more components each individually selected from the group consisting of an amino acid, an amino acid residue, an amino acid analog, and a modified amino acid. The components are typically joined to each other through an amide bond.
  • the peptide such as the antibody, is PEGylated.
  • PEGylation can provide, for example, increased stability and/or efficacy of the polypeptide.
  • Methods for PEGylation known in the art can be used in the methods and compositions provided herein.
  • Such methods include, but are not limited to, those provided in “Comparative Binding of Disulfide-Bridged PEG-Fabs” Khalili et al., Bioconjugate Chemistry (2012), 23(11), 2262-2277; “Site-Specific PEGylation at Histidine Tags” Cong et al., Bioconjugate Chemistry (2012), 23(2), 248-263; “Disulfide bridge based PEGylation of proteins” Brocchini et al., Advanced Drug Delivery Reviews (2008), 60(1), 3-12; “Site-Specific PEGylation of Protein Disulfide Bonds Using a Three-Carbon Bridge” Balan et al., Bioconjugate Chemistry (2007), 18(1), 61-76; “Site-specific PEGylation of native disulfide bonds in therapeutic proteins” Shaunak et al., Nature Chemical Biology (2006), 2(6), 312-313; “PEG derivative conjugated proteins and peptides for use in pharmaceuticals” Godwin et al
  • amino acid includes naturally occurring amino acids, a molecule having a nitrogen available for forming an amide bond and a carboxylic acid, a molecule of the general formula NH 2 —CHR—COOH or the residue within a peptide bearing the parent amino acid, where “R” is one of a number of different side chains. “R” can be a substituent found in naturally occurring amino acids. “R” can also be a substituent referring to one that is not of the naturally occurring amino acids.
  • amino acid residue refers to the portion of the amino acid which remains after losing a water molecule when it is joined to another amino acid.
  • amino acid analog refers to a structural derivative of an amino acid parent compound that often differs from it by a single element.
  • modified amino acid refers to an amino acid bearing an “R” substituent that does not correspond to one of the twenty genetically coded amino acids.
  • the abbreviations for the genetically encoded L-enantiomeric amino acids are conventional and are as follows:
  • amino acid residues in the active agent-conjugate can be replaced with other amino acid residues without significantly deleteriously affecting, and in many cases even enhancing, the activity of the peptides.
  • altered or mutated forms of the active agent-conjugate wherein at least one defined amino acid residue in the structure is substituted with another amino acid residue or derivative and/or analog thereof. It will be recognized that in preferred embodiments, the amino acid substitutions are conservative, i.e., the replacing amino acid residue has physical and chemical properties that are similar to the amino acid residue being replaced.
  • the amino acids can be conveniently classified into two main categories—hydrophilic and hydrophobic—depending primarily on the physical-chemical characteristics of the amino acid side chain. These two main categories can be further classified into subcategories that more distinctly define the characteristics of the amino acid side chains.
  • hydrophilic amino acids can be further subdivided into acidic, basic and polar amino acids.
  • hydrophobic amino acids can be further subdivided into nonpolar and aromatic amino acids.
  • hydrophilic amino acid refers to an amino acid exhibiting a hydrophobicity of less than zero according to the normalized consensus hydrophobicity scale of Eisenberg et al., 1984, J. Mol. Biol. 179:125-142. Genetically encoded hydrophilic amino acids include Thr (T), Ser (S), His (H), Glu (E), Asn (N), Gln (Q), Asp (D), Lys (K) and Arg (R).
  • hydrophobic amino acid refers to an amino acid exhibiting a hydrophobicity of greater than zero according to the normalized consensus hydrophobicity scale of Eisenberg, 1984, J. Mol. Biol. 179:1.25-142. Genetically encoded hydrophobic amino acids include Pro (P), Ile (I), Phe (F), Val (V), Leu (L), Trp (W), Met (M), Ala (A), Gly (G) and Tyr (Y).
  • acidic amino acid refers to a hydrophilic amino acid having a side chain pK value of less than 7. Acidic amino acids typically have negatively charged side chains at physiological pH due to loss of a hydrogen ion. Genetically encoded acidic amino acids include Glu (E) and Asp (D).
  • basic amino acid refers to a hydrophilic amino acid having a side chain pK value of greater than 7.
  • Basic amino acids typically have positively charged side chains at physiological pH due to association with hydronium ion.
  • Genetically encoded basic amino acids include His (H), Arg (R) and Lys (K).
  • polar amino acid refers to a hydrophilic amino acid having a side chain that is uncharged at physiological pH, but which has at least one bond in which the pair of electrons shared in common by two atoms is held more closely by one of the atoms.
  • Genetically encoded polar amino acids include Asn (N), Gln (Q) Ser (S) and Thr (T).
  • nonpolar amino acid refers to a hydrophobic amino acid having a side chain that is uncharged at physiological pH and which has bonds in which the pair of electrons shared in common by two atoms is generally held equally by each of the two atoms (i.e., the side chain is not polar).
  • Genetically encoded nonpolar amino acids include Leu (L), Val (V), Ile (I), Met (M), Gly (G) and Ala (A).
  • aromatic amino acid refers to a hydrophobic amino acid with a side chain having at least one aromatic or heteroaromatic ring.
  • the aromatic or heteroaromatic ring may contain one or more substituents such as OH, SH, CN, F, Cl, Br, I, NO 2 , NO, NH 2 , NHR, NRR, —C(O)R, —C(O)OH, —C(O)OR, —C(O)NH 2 , —C(O)NHR, —C(O)NRR and the like where each R is independently (C 1 -C 6 ) alkyl, substituted (C 1 -C 6 ) alkyl, (C 1 -C 6 ) alkenyl, substituted (C 1 -C 6 ) alkenyl, (C 1 -C 6 ) alkynyl, substituted (C 1 -C 6 ) alkynyl, (C 5 -C 20 ) aryl
  • aliphatic amino acid refers to a hydrophobic amino acid having an aliphatic hydrocarbon side chain. Genetically encoded aliphatic amino acids include Ala (A), Val (V), Leu (L) and Ile (I).
  • Cys (C) is unusual in that it can form disulfide bridges with other Cys (C) residues or other sulfanyl-containing amino acids.
  • the ability of Cys (C) residues (and other amino acids with —SH containing side chains) to exist in a peptide in either the reduced free —SH or oxidized disulfide-bridged form affects whether Cys (C) residues contribute net hydrophobic or hydrophilic character to a peptide.
  • Cys (C) exhibits a hydrophobicity of 0.29 according to the normalized consensus scale of Eisenberg (Eisenberg, 1984, supra), it is to be understood that for purposes of the preferred embodiments Cys (C) is categorized as a polar hydrophilic amino acid, notwithstanding the general classifications defined above.
  • amino acids having side chains exhibiting two or more physical-chemical properties can be included in multiple categories.
  • amino acid side chains having aromatic moieties that are further substituted with polar substituents, such as Tyr (Y) may exhibit both aromatic hydrophobic properties and polar or hydrophilic properties, and can therefore be included in both the aromatic and polar categories.
  • polar substituents such as Tyr (Y)
  • the amino acid substitutions need not be, and in certain embodiments preferably are not, restricted to the genetically encoded amino acids.
  • the active agent-conjugate may contain genetically non-encoded amino acids.
  • amino acid residues in the active agent-conjugate may be substituted with naturally occurring non-encoded amino acids and synthetic amino acids.
  • Certain commonly encountered amino acids which provide useful substitutions for the active agent-conjugates include, but are not limited to, ⁇ -alanine ( ⁇ -Ala) and other omega-amino acids such as 3-aminopropionic acid, 2,3-diaminopropionic acid (Dpr), 4-aminobutyric acid and so forth; ⁇ -aminoisobutyric acid (Aib); ⁇ -aminohexanoic acid (Aha); ⁇ -aminovaleric acid (Ava); N-methylglycine or sarcosine (MeGly); ornithine (Orn); citrullinc (Cit); t-butylalaninc (t-BuA); t-butylglycinc (t-BuG); N-methylisoleucine (Melle); phenylglycine (Phg); cyclohexylalanine (Cha); norleucine (Nle); naphthylalanine (Nal
  • amino acid residues not specifically mentioned herein can be readily categorized based on their observed physical and chemical properties in light of the definitions provided herein.
  • Table 2 The classifications of the genetically encoded and common non-encoded amino acids according to the categories defined above are summarized in Table 2, below. It is to be understood that Table 2 is for illustrative purposes only and does not purport to be an exhaustive list of amino acid residues and derivatives that can be used to substitute the active agent-conjugate described herein.
  • amino acid residues not specifically mentioned herein can be readily categorized based on their observed physical and chemical properties in light of the definitions provided herein.
  • the amino acids of the active agent-conjugate will be substituted with L-enantiomeric amino acids, the substitutions are not limited to L-enantiomeric amino acids.
  • the peptides may advantageously be composed of at least one D-enantiomeric amino acid. Peptides containing such D-amino acids are thought to be more stable to degradation in the oral cavity, gut or serum than are peptides composed exclusively of L-amino acids.
  • Examples include, but are not limited:
  • Examples include, but are not limited, the following:
  • the resultant crude ADC product underwent necessary down-stream steps of desalt, buffet changes/formulation, and optionally, purification, using the state-of-art procedures.
  • the final ADC product was characterized by HIC-HPLC, SEC, RP-HPLC, and optionally LC-MS.
  • the average DAR was calculated by UV absorption and/or MS spectroscopy.
  • the acid sensitive protecting groups containing compound was dissolved in 4N HCl/dioxane and the mixture was stirred at room temperature for 2 h. The solution was then concentrated under reduced pressure and the residue was washed twice with cold ether. Purification was carried out on reverse phase HPLC if necessary.
  • the Fmoc containing compound was dissolved in 2-5% piperidine in DMF. The mixture was stirred at room temperature for 1 h. The solvents were removed under reduced pressure. Purification was carried out on reverse phase HPLC if necessary.
  • Method 1-1 Linker and Drug Connected Via a carbamate Bond Using General Procedure G or I for Activation and carbamate Formation and General Procedure C, D, or F for Removal of Protective Groups.
  • Method 1-3 Active molecule containing a carboxylic acid moiety connected to an alkoxyamino linker via formation of hydoxamate (General procedure A or B), followed by removal of protective groups.
  • Step 3 Introducing the final functional groups prior to conjugation
  • amino acid sulfonamide derivatives 11 were synthesized according to previously reported procedure (ARKIVOC 2004 (xii) 14-22, or WO 2007146695) using Boc protected amino acid and cyclopropyl/methyl sulfonamide, followed by removal of Boc (General procedure C)
  • amino acid sulfonamide derivatives 14 were synthesized according to previously reported procedure (ARKIVOC 2004 (xii) 14-22, or WO 2007146695) using Boc protected amino acid and cyclopropyl/methyl sulfonamide, followed by removal of Boc (General procedure C).
  • Compound 16 was synthesized using the general procedures described above as following: DIC/HOAt mediated amide bond formation (General procedure B) between Boc-N-Me-Val-Val-Dil-Dap-OH (compound 10) and amine 14, followed by removal of Boc (General procedure C). The final compound was purified by reverse phase HPLC to give compound 16 as a white powder after lyophilization. MS m/z Calcd for C 41 H 70 N 6 O 10 S 838.5, Found 839.6 ([M+H] + ).
  • Compound 32 was synthesized using the general procedures described above as following: DIC/HOAt mediated amide bond formation (General procedure B) between Auristatin F and compound 26, followed by removal of Fmoc (General procedure D), reaction with acid 30 (General procedure A), and reduction of the nitro group to amino group.
  • nitro compound 31 (32 mg) was dissolved in acetonitrile/water (6/4, v/v) and a solution of sodium dithionite (1M in water, 0.2 mL) was added. The pH of the reaction was controlled at 6-6.5 by addition of MES buffer. The mixture was stirred at room temperature for 2 h and purified directly by RP-HPLC to give the desired product as a white solid after lyophilization (18 mg). MS found 1319.0 (M+H) + .
  • the o-phenylenediamine compound 32 (12 mg) was dissolved in acetonitrile/water (1 mL). Dibromomethyl diketone (10 mg) was added. The mixture was stirred at room temperature and purified directly by RP-HPLC to give the desired quinoxaline 40 as a white powder (12 mg) after lyophilization. MS found 1526.4 (M+H) + .
  • Fmoc-Ahx-OH (353 mg) was suspended in anhydrous DCM (5 mL) and DIC (0.5 mmol) was added. The mixture was stirred at room temperature for 1 h. Maytansinol (compound 51, 56 mg) was added to the mixture, followed by addition of 0.5 mL of pyridine. The reaction mixture was stirred at room temperature for 48 h.
  • the resultant crude ADC product underwent necessary down-stream steps of desalt, buffet changes/formulation, and optionally, purification, using the state-of-art procedures.
  • the final ADC product was characterized by HIC-HPLC, SEC, RP-HPLC, and optionally LC-MS.
  • the average DAR was calculated by UV absorption and/or MS spectroscopy.
  • Trastuzumab and compound 34 were combined in various drug/antibody ratios under as described in the General Conjugation Procedure.
  • Trastuzumab was incubated with 10 mM TCEP stock solution at 37° C. for 0.5 to 40 hours. After reduction, the excess of TCEP was removed by a gel filtration column. The reduced trastuzumab was then mixed with compound 34 in drug/antibody ratio range from 1 to 10 at 0 to 40° C. After overnight reaction, excess compound 34 was removed from Trastuzumab-c-34 conjugate by a gel filtration column. Purified Trastuzumab-c-34 conjugates stored at 4 to ⁇ 20° C. and to be used for in vitro and in vivo tests.
  • the product of the conjugation reaction was analyzed using HIC-HPLC under conditions: HPLC Column: Tosoh TSKgel Butyl-NPR, 4.6 mm ⁇ 3.5 cm, 2.5 mm; Buffer A: 20 mM sodium phosphate, 1.5 M Ammonium Sulfate, pH 7.0; Buffer B: 20 mM sodium phosphate, 25% v/v isopronal, pH 7.0; flow rate: 1 mL/min; Gradient: 10 min 10% Buffer B to 80% Buffer B, 4 min 100% Buffer B; 20 ⁇ L sample.
  • FIG. 1 shows the HIC-HPLC chromatogram product of reacting drug and antibody at a drug/antibody ratio of 5.5:1, where DAR represents the number of drugs conjugated per antibody.
  • FIG. 2 shows the HIC-HPLC chromatogram product of reacting drug and antibody at a drug/antibody ratio of 6:1, where DAR represents the number of drugs conjugated per antibody.
  • FIG. 3 shows the HIC-HPLC chromatogram product of reacting drug and antibody at a drug/antibody ratio of 1:6.5, where DAR represents the number of drugs conjugated per antibody.
  • FIG. 4 shows the HIC-HPLC chromatogram product of reacting drug and antibody at a drug/antibody ratio of 1:4, where DAR represents the number of drugs conjugated per antibody.
  • FIG. 4 shows the HIC-HPLC chromatogram product of reacting drug and antibody at a drug/antibody ratio of 1:4, where DAR represents the number of drugs conjugated per antibody.
  • FIG. 4 shows the HIC-HPLC chromatogram product of reacting
  • FIG. 4 further shows HIC-HPLC chromatograms for individually isolated fractions of DAR 1, DAR 2, DAR 3, DAR 4 and DAR 5 peaks purified using FPLC.
  • FIG. 5 shows HIC-HPLC chromatograms for each FPLC fraction, and, inset, an overlay of the peaks to demonstrate relative purity of each peak.
  • FIGS. 4 and 5 The purified fractions depicted in FIGS. 4 and 5 were further analyzed by reduced SDS-PAGE.
  • the ADC samples were reduced with 20 mM DTT and loaded onto a NuPAGE 4-12% Bis-Tris Gel (Life Technology). The results are shown in FIG. 6 , where the abbreviations are as follows:
  • the antibody drug conjugates were the analyzed for cytotoxicity.
  • the cell lines used were SK-BR-3 human breast adenocarcinoma (HER2 triple positive), HCC1954 human Ductal Carcinoma (HER2 triple positive). These cells were available from ATCC.
  • SK-BR-3 cells were grown in McCoy's 5A medium (Caisson Labs, North Logan, UT) supplemented with 10% fetal bovine serum.
  • HCC1954 cells were grown in RPMI-1640 medium (Caisson Labs, North Logan, UT) supplemented with 10% fetal bovine serum.
  • SK-BR-3 cells were plated in 96-well plates at approximately 7,500 cells/well, and HCC1954 cells were plated in 96-well plates at approximately 20,000 cells/well.
  • FIG. 7 shows viability of SK-BR-3 cells for seven samples, ADC_C1 (DAR1 from FIGS. 4 and 5 ), ADC_C2 (DAR2 from FIGS. 4 and 5 ), ADC_C3 (DAR3 from FIGS. 4 and 5 ), ADC_C4 (DAR4 from FIGS. 4 and 5 ), ADC_5 (DAR5 from FIGS. 4 and 5 ), SeaGen (ADC prepared following published Seattle Genetics method), and T-DM1 (ADC prepared following published Immunogen method).
  • ADC_C1 DAR1 from FIGS. 4 and 5
  • ADC_C2 DAR2 from FIGS. 4 and 5
  • ADC_C3 DAR3 from FIGS. 4 and 5
  • ADC_C4 DAR4 from FIGS. 4 and 5
  • ADC_5 DAR5 from FIGS. 4 and 5
  • SeaGen ADC prepared following published Seattle Genetics method
  • T-DM1 ADC prepared following published Immunogen method
  • FIG. 8 shows viability of HCC1954 cells for seven samples, ADC_C1 (DAR1 from FIGS. 4 and 5 ), ADC_C2 (DAR2 from FIGS. 4 and 5 ), ADC_C3 (DAR3 from FIGS. 4 and 5 ), ADC_C4 (DAR4 from FIGS. 4 and 5 ), ADC_C5 (DAR5 from FIGS. 4 and 5 ), ADC_VC-M MAE was prepared following published Seattle Genetics method), and T-DM1 (ADC prepared following published Immunogen method).
  • FIG. 10 shows viability of SK-BR-3 cells for four samples, both ADC_C6 (DAR2.4) and ADC_C7 (DAR3.1) were prepared using the General Conjugation Procedure and purified by a gel filtration column. SeaGen (ADC prepared following published Seattle Genetics method), and T-DM1 (ADC prepared following published Immunogen method).
  • FIG. 11 shows viability of HCC1954 cells for four samples, ADC_C6 (DAR2.4 from FIG. 10 ), ADC_C7 (DAR3.1 from FIG. 10 ), ADC_VC-M MAE was prepared following published Seattle Genetics method), and T-DM1 (ADC prepared following published Immunogen method).
  • Trastuzumab and compound 42 were combined in various drug/antibody ratios under conditions where Trastuzumab was incubated with 10 mM TCEP stock solution at 37° C. for 0.5 to 40 hours. After reduction, the excess of TCEP was removed by a gel filtration column. The reduced Trastuzumab was then mixed with compound 42 in drug/antibody ratio range from 1 to 10 at 0 to 40° C. After overnight reaction, excess compound 42 was removed from Trastuzumab-c-34 conjugate by a gel filtration column. Purified Trastuzumab-c-42 conjugates stored at 4 to ⁇ 20° C. and to be used for in vitro and in vivo tests.
  • Trastuzumab and compound 40 were combined in various drug/antibody ratios under conditions where Trastuzumab was incubated with 10 mM TCEP stock solution at 37° C. for 0.5 to 40 hours. After reduction, the excess of TCEP was removed by a gel filtration column. The reduced Trastuzumab was then mixed with compound 40 in drug/antibody ratio range from 1 to 10 at 0 to 40° C. After overnight reaction, excess compound 40 was removed from Trastuzumab-c-34 conjugate by a gel filtration column. Purified Trastuzumab-c-40 conjugates stored at 4 to ⁇ 20° C. and to be used for in vitro and in vivo tests.
  • the product of the conjugation reaction was analyzed using reduced SDS-PAGE.
  • the ADC samples were reduced with 20 mM DTT and loaded onto a NuPAGE 4-12% Bis-Tris Gel (Life Technology). The results are shown in FIG. 9 , where the abbreviations are as follows:
  • the Trastuzumab-compound 42 and Trastuzumab-compound 42 conjugates were the analyzed for cytotoxicity.
  • the cytotoxicity methods were performed as described above.
  • FIG. 12 shows viability of SK-BR-3 cells for four samples, Trastuzumab-c-Duo3 (Trastuzumab-compound 42 conjugate), Trastuzumab-c-Duo6 (Trastuzumab-compound 40 conjugate), Trastuzumab-vc-MMAE (prepared following published Seattle Genetics method), and Trastuzumab-c-MMAE (Trastuzumab-compound 34 conjugate, prepared following published Seattle Genetics method).

Landscapes

  • Health & Medical Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Organic Chemistry (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Engineering & Computer Science (AREA)
  • Medicinal Chemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Veterinary Medicine (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Animal Behavior & Ethology (AREA)
  • Public Health (AREA)
  • Immunology (AREA)
  • Epidemiology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Molecular Biology (AREA)
  • Genetics & Genomics (AREA)
  • Biochemistry (AREA)
  • Biophysics (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • General Chemical & Material Sciences (AREA)
  • Cell Biology (AREA)
  • Diabetes (AREA)
  • Oncology (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Obesity (AREA)
  • Hematology (AREA)
  • Communicable Diseases (AREA)
  • Endocrinology (AREA)
  • Emergency Medicine (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
  • Peptides Or Proteins (AREA)
  • Medicinal Preparation (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
  • Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)

Abstract

In one aspect, an active agent-conjugate, methods of preparing the active agent-conjugate, and uses thereof is provided.

Description

RELATED APPLICATIONS
This application is a continuation of U.S. application Ser. No. 15/854,742, filed on Dec. 26, 2017, which is a continuation of U.S. application Ser. No. 14/401,114, filed on Nov. 13, 2014 and issued as U.S. Pat. No. 9,884,127 on Feb. 6, 2018, which is a 35 U.S.C. § 371 national stage of International Application No. PCT/US2013/041026, filed on May 14, 2013. PCT/US2013/041026 claims the benefit of U.S. Provisional Patent Application No. 61/652,512, filed on May 29, 2012, U.S. Provisional Patent Application No. 61/648,406, filed on May 17, 2012, U.S. Provisional Patent Application No. 61/648,532, filed on May 17, 2012, and U.S. Provisional Patent Application No. 61/647,300, filed on May 15, 2012. The entire contents of each of the aforementioned applications are hereby incorporated by reference.
BACKGROUND
Recently, it has been found that an antibody (or antibody fragment such as a single-chain variable fragment) can be linked to a payload drug to form an immunoconjugate that has been termed antibody-drug conjugate, or ADC. The antibody causes the ADC to bind to the target cells. Often the ADC is then internalized by the cell and the drug is released to treat the cell. Because of the targeting, the side effects may be lower than the side effects of systemically administering the drug.
SUMMARY
Some embodiments provide active agent-conjugates, methods of preparing active agent-conjugates, and uses thereof.
Some embodiments provide an active agent-conjugate having the structure of Formula I:
Figure US12115228-20241015-C00001
    • or a pharmaceutically acceptable salt thereof,
    • wherein:
    • A may be a targeting moiety;
    • B may be an auxiliary moiety that optionally is a second targeting moiety, or B may be null;
    • L1 includes a 2- to 5-carbon bridge and at least one sulfur atom;
    • each D may be independently selected, where each D includes an active agent;
    • each L2 may be independently a linker, wherein at least one L2 links to L1; and
    • n may be 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.
Some embodiments provide an active agent-conjugate having the structure of Formula Ia:
Figure US12115228-20241015-C00002
    • or a pharmaceutically acceptable salt thereof,
    • wherein:
    • the A-component may be a targeting moiety;
    • the E-component may be an optionally substituted heteroaryl or an optionally substituted heterocyclyl;
    • B may be an auxiliary moiety that optionally is a second targeting moiety, or B may be null;
    • each D may be independently selected, where each D includes an active agent;
    • each L2 may be independently a linker, wherein at least one L2 links to L1; and
    • n may be 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10;
    • L3 may be an optionally substituted C1-C6 alkyl, or L3 may be null, when L3 is null the sulfur is directly connected to the E-component; and
    • L4 may be an optionally substituted C1-C6 alkyl, or L4 may be null, when L4 is null the sulfur is directly connected to the E-component.
In some embodiments, the E-component includes a fragment selected from the group consisting of:
Figure US12115228-20241015-C00003
In some embodiments, L3 may be —(CH2)—; and L4 may be —(CH2)—. In some embodiments, L3 may be null; and L4 may be null.
In some embodiments, be:
Figure US12115228-20241015-C00004
In some embodiments, L2 includes —(CH2)n— where n may be 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. In some embodiments, L2 includes —(CH2CH2O)n— where n may be 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. In some embodiments, L2 includes Val-Cit-PAB, Val-Ala-PAB, Val-Lys(Ac)-PAB, Phe-Lys-PAB, Phe-Lys(Ac)-PAB, D-Val-Leu-Lys, Gly-Gly-Arg, Ala-Ala-Asn-PAB, Ala-PAB, or PAB. In some embodiments, L2 includes peptide, oligosaccharide, —(CH2)n—, —(CH2CH2O)n—, Val-Cit-PAB, Val-Ala-PAB, Val-Lys(Ac)-PAB, Phe-Lys-PAB, Phe-Lys(Ac)-PAB, D-Val-Leu-Lys, Gly-Gly-Arg, Ala-Ala-Asn-PAB, Ala-PAB, PAB, or combinations thereof. In some embodiments, L2 includes a noncleavable unit. In some embodiments, the noncleavable unit includes —(CH2)n— where n may be 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. In some embodiments, the noncleavable unit includes —(CH2CH2O)n— where n may be 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. In some embodiments, L2 includes a cleavable unit. In some embodiments, the cleavable unit comprises a peptide.
In some embodiments, the A component comprises a monoclonal antibody (mAB). In some embodiments, the A component comprises an antibody fragment, surrogate, or variant. In some embodiments, the A component comprises a protein ligand. In some embodiments, the A component comprises a protein scaffold. In some embodiments, the A component comprises a peptide. In some embodiments, the A component comprises a small molecule ligand. In some embodiments, the A component comprises at least one modified L-Alanine residue. In some embodiments, the A component comprises at least two modified L-Alanine residues. In some embodiments, at least one L2 includes —(CH2)n— where n may be 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. In some embodiments, at least one L2 includes —(CH2CH2O)n— where n may be 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. In some embodiments, at least one L2 includes Val-Cit-PAB, Val-Ala-PAB, Val-Lys(Ac)-PAB, Phe-Lys-PAB, Phe-Lys(Ac)-PAB, D-Val-Leu-Lys, Gly-Gly-Arg, Ala-Ala-Asn-PAB, Ala-PAB, or PAB. In some embodiments, at least one L2 includes a peptide, an oligosaccharide, —(CH2)n—, —(CH2CH2O)n—, Val-Cit-PAB, Val-Ala-PAB, Val-Lys(Ac)-PAB, Phe-Lys-PAB, Phe-Lys(Ac)-PAB, D-Val-Leu-Lys, Gly-Gly-Arg, Ala-Ala-Asn-PAB, Ala-PAB, PAB or combinations thereof.
BRIEF DESCRIPTION OF THE DRAWINGS
FIG. 1 shows the HIC-HPLC chromatogram product of reacting drug and antibody at a drug/antibody ratio of 5.5:1.
FIG. 2 shows the HIC-HPLC chromatogram product of reacting drug and antibody at a drug/antibody ratio of 6:1.
FIG. 3 shows the HIC-HPLC chromatogram product of reacting drug and antibody at a drug/antibody ratio of 1:6.5.
FIG. 4 shows the HIC-HPLC chromatogram product of reacting drug and antibody at a drug/antibody ratio of 1:4
FIG. 5 shows HIC-HPLC chromatograms for each FPLC fraction, and, inset, an overlay of the peaks to demonstrate relative purity of each peak.
FIG. 6 shows reduced SDS-PAGE analysis for the FPLC fractions depicted in FIGS. 4 and 5 .
FIG. 7 shows viability of SK-BR-3 cells for seven antibody-drug conjugate samples.
FIG. 8 shows viability of HCC1954 cells for seven antibody-drug conjugate samples.
FIG. 9 shows reduced SDS-PAGE analysis for trastuzumab conjugated compound 40.
FIG. 10 shows viability of SK-BR-3 cells for four antibody-drug conjugate samples.
FIG. 11 shows viability of HCC1954 cells for four antibody-drug conjugate samples.
FIG. 12 shows viability of SK-BR-3 cells for four antibody-drug conjugate samples.
 DETAILED DESCRIPTION
Some embodiments provide an active agent-conjugate. In some embodiments, the active agent-conjugate is a drug-conjugate. In some embodiments, the drug-conjugate includes a targeting molecule. In some embodiments, the targeting molecule includes a monoclonal antibody (mAB). In some embodiments, the drug-conjugate includes a spacer or a multifunctional linker. In some embodiments, the spacer connects to the mAB by a sulfide bond. In some embodiments, the multifunctional linker connects to the mAB by a sulfide bond. In some embodiments, the spacer or multifunctional linker may be optionally connected to an auxiliary moiety. In some embodiments, the auxiliary moiety may be a second targeting molecule such as mAB and peptide. In some embodiments, the auxiliary moiety may be a hydrophilic polymer such as polyethylene glycol (PEG), and the like. In some embodiments, the spacer or multifunctional linker may include a 2- to 5-atom bridge. In some embodiments, the spacer or multifunctional linker may include a 4C (four carbon) bridge.
Conjugation methods to derivatize a polypeptide with a payload can be accomplished using a maleimido or vinyl moiety which can react with individual sulfhydryl group on an antibody via Michael addition reaction. The free sulfhydryl group can be formed by reducing a disulfide bond in an antibody. However, the structural integrity of an antibody can be compromised after opening disulfide bonds and attaching payloads to the exposed free thiols. The compositions and methods provided herein provide conjugation through cysteine without decreased structural stability.
Definitions
As used herein, common organic abbreviations are defined as follows:
    • Ac Acetyl
    • aq. Aqueous
    • BOC or Boc tert-Butoxycarbonyl
    • BrOP bromo tris(dimethylamino) phosphonium hexafluorophosphate
    • Bu n-Butyl
    • ° C. Temperature in degrees Centigrade
    • DCM methylene chloride
    • DEPC Diethylcyanophosphonate
    • DIC diisopropylcarbodiimide
    • DIEA Diisopropylethylamine
    • DMA N,N′-Dimethylacetamide
    • DMF N,N′-Dimethylformamide
    • DTT Dithiothreitol
    • EDC 1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide
    • Et Ethyl
    • EtOAc Ethyl acetate
    • Eq Equivalents
    • Fmoc 9-Fluorenylmethoxycarbonyl
    • g Gram(s)
    • h Hour (hours)
    • HATU 2-(1H-7-azabenzotriazol-1-yl)-1,1,3,3-tetramethyl uronium hexafluorophosphate
    • HOAt 1-Hydroxy-7-azabenzotriazole
    • HOBT N-Hydroxybenzotriazole
    • HPLC High-performance liquid chromatography
    • LC/MS Liquid chromatography-mass spectrometry
    • Me Methyl
    • MeOH Methanol
    • MeCN Acetonitrile
    • mL Milliliter(s)
    • MS mass spectrometry
    • PAB p-aminobenzyl
    • RP-HPLC reverse phase HPLC
    • t-Bu tert-Butyl
    • TCEP Tris(2-carboxyethyl)phosphine
    • TEA Triethylamine
    • Tert, t tertiary
    • TFA Trifluoracetic acid
    • THF Tetrahydrofuran
    • THP Tetrahydropyranyl
    • TLC Thin-layer chromatography
    • μL Microliter(s)
The term “pharmaceutically acceptable salt” refers to salts that retain the biological effectiveness and properties of a compound and, which are not biologically or otherwise undesirable for use in a pharmaceutical. In many cases, the compounds disclosed herein are capable of forming acid and/or base salts by virtue of the presence of amino and/or carboxyl groups or groups similar thereto. Pharmaceutically acceptable acid addition salts can be formed with inorganic acids and organic acids. Inorganic acids from which salts can be derived include, for example, hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, and the like. Organic acids from which salts can be derived include, for example, acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, salicylic acid, and the like. Pharmaceutically acceptable base addition salts can be formed with inorganic and organic bases. Inorganic bases from which salts can be derived include, for example, sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese, aluminum, and the like; particularly preferred are the ammonium, potassium, sodium, calcium and magnesium salts. Organic bases from which salts can be derived include, for example, primary, secondary, and tertiary amines, substituted amines including naturally occurring substituted amines, cyclic amines, basic ion exchange resins, and the like, specifically such as isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, and ethanolamine. Many such salts are known in the art, as described in WO 87/05297, Johnston et al., published Sep. 11, 1987 (incorporated by reference herein in its entirety).
As used herein, “Ca to Cb ” or “Ca-b” in which “a” and “b” are integers refer to the number of carbon atoms in the specified group. That is, the group can contain from “a” to “b”, inclusive, carbon atoms. Thus, for example, a “C1 to C4 alkyl” or “C1-4 alkyl” group refers to all alkyl groups having from 1 to 4 carbons, that is, CH3—, CH3CH2—, CH3CH2CH2—, (CH3)2CH—, CH3CH2CH2CH2—, CH3CH2CH(CH3)— and (CH3)3C—.
The term “halogen” or “halo,” as used herein, means any one of the radio-stable atoms of column 7 of the Periodic Table of the Elements, e.g., fluorine, chlorine, bromine, or iodine, with fluorine and chlorine being preferred.
As used herein, “alkyl” refers to a straight or branched hydrocarbon chain that is fully saturated (i.e., contains no double or triple bonds). The alkyl group may have 1 to 20 carbon atoms (whenever it appears herein, a numerical range such as “1 to 20” refers to each integer in the given range; e.g., “1 to 20 carbon atoms” means that the alkyl group may consist of 1 carbon atom, 2 carbon atoms, 3 carbon atoms, etc., up to and including 20 carbon atoms, although the present definition also covers the occurrence of the term “alkyl” where no numerical range is designated). The alkyl group may also be a medium size alkyl having 1 to 9 carbon atoms. The alkyl group could also be a lower alkyl having 1 to 4 carbon atoms. The alkyl group may be designated as “C1-4 alkyl” or similar designations. By way of example only, “C1-4 alkyl” indicates that there are one to four carbon atoms in the alkyl chain, i.e., the alkyl chain is selected from the group consisting of methyl, ethyl, propyl, iso-propyl, n-butyl, iso-butyl, sec-butyl, and t-butyl. Typical alkyl groups include, but are in no way limited to, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, tertiary butyl, pentyl, hexyl, and the like.
As used herein, “alkoxy” refers to the formula —OR wherein R is an alkyl as is defined above, such as “C1-9 alkoxy”, including but not limited to methoxy, ethoxy, n-propoxy, 1-methylethoxy (isopropoxy), n-butoxy, iso-butoxy, sec-butoxy, and tert-butoxy, and the like.
As used herein, “alkylthio” refers to the formula —SR wherein R is an alkyl as is defined above, such as “C1-9 alkylthio” and the like, including but not limited to methylmercapto, ethylmercapto, n-propylmercapto, 1-methylethylmercapto (isopropylmercapto), n-butylmercapto, iso-butylmercapto, sec-butylmercapto, tert-butylmercapto, and the like.
As used herein, “alkenyl” refers to a straight or branched hydrocarbon chain containing one or more double bonds. The alkenyl group may have 2 to 20 carbon atoms, although the present definition also covers the occurrence of the term “alkenyl” where no numerical range is designated. The alkenyl group may also be a medium size alkenyl having 2 to 9 carbon atoms. The alkenyl group could also be a lower alkenyl having 2 to 4 carbon atoms. The alkenyl group may be designated as “C2-4 alkenyl” or similar designations. By way of example only, “C2-4 alkenyl” indicates that there are two to four carbon atoms in the alkenyl chain, i.e., the alkenyl chain is selected from the group consisting of ethenyl, propen-1-yl, propen-2-yl, propen-3-yl, buten-1-yl, buten-2-yl, buten-3-yl, buten-4-yl, 1-methyl-propen-1-yl, 2-methyl-propen-1-yl, 1-ethyl-ethen-1-yl, 2-methyl-propen-3-yl, buta-1,3-dienyl, buta-1,2,-dienyl, and buta-1,2-dien-4-yl. Typical alkenyl groups include, but are in no way limited to, ethenyl, propenyl, butenyl, pentenyl, and hexenyl, and the like.
As used herein, “alkynyl” refers to a straight or branched hydrocarbon chain containing one or more triple bonds. The alkynyl group may have 2 to 20 carbon atoms, although the present definition also covers the occurrence of the term “alkynyl” where no numerical range is designated. The alkynyl group may also be a medium size alkynyl having 2 to 9 carbon atoms. The alkynyl group could also be a lower alkynyl having 2 to 4 carbon atoms. The alkynyl group may be designated as “C2-4 alkynyl” or similar designations. By way of example only, “C2-4 alkynyl” indicates that there are two to four carbon atoms in the alkynyl chain, i.e., the alkynyl chain is selected from the group consisting of ethynyl, propyn-1-yl, propyn-2-yl, butyn-1-yl, butyn-3-yl, butyn-4-yl, and 2-butynyl. Typical alkynyl groups include, but are in no way limited to, ethynyl, propynyl, butynyl, pentynyl, and hexynyl, and the like.
The term “aromatic” refers to a ring or ring system having a conjugated pi electron system and includes both carbocyclic aromatic (e.g., phenyl) and heterocyclic aromatic groups (e.g., pyridine). The term includes monocyclic or fused-ring polycyclic (i.e., rings which share adjacent pairs of atoms) groups provided that the entire ring system is aromatic.
As used herein, “aryloxy” and “arylthio” refers to RO- and RS-, in which R is an aryl as is defined above, such as “C6-10 aryloxy” or “C6-10 arylthio” and the like, including but not limited to phenyloxy.
An “aralkyl” or “arylalkyl” is an aryl group connected, as a substituent, via an alkylene group, such as “C7-14 aralkyl” and the like, including but not limited to benzyl, 2-phenylethyl, 3-phenylpropyl, and naphthylalkyl. In some cases, the alkylene group is a lower alkylene group (i.e., a C1-4 alkylene group).
As used herein, “heteroaryl” refers to an aromatic ring or ring system (i.e., two or more fused rings that share two adjacent atoms) that contain(s) one or more heteroatoms, that is, an element other than carbon, including but not limited to, nitrogen, oxygen and sulfur, in the ring backbone. When the heteroaryl is a ring system, every ring in the system is aromatic. The heteroaryl group may have 5-18 ring members (i.e., the number of atoms making up the ring backbone, including carbon atoms and heteroatoms), although the present definition also covers the occurrence of the term “heteroaryl” where no numerical range is designated. In some embodiments, the heteroaryl group has 5 to 10 ring members or 5 to 7 ring members. The heteroaryl group may be designated as “5-7 membered heteroaryl,” “5-10 membered heteroaryl,” or similar designations. Examples of heteroaryl rings include, but are not limited to, furyl, thienyl, phthalazinyl, pyrrolyl, oxazolyl, thiazolyl, imidazolyl, pyrazolyl, isoxazolyl, isothiazolyl, triazolyl, thiadiazolyl, pyridinyl, pyridazinyl, pyrimidinyl, pyrazinyl, triazinyl, quinolinyl, isoquinlinyl, benzimidazolyl, benzoxazolyl, benzothiazolyl, indolyl, isoindolyl, and benzothienyl.
A “heteroaralkyl” or “heteroarylalkyl” is heteroaryl group connected, as a substituent, via an alkylene group. Examples include but are not limited to 2-thienylmethyl, 3-thienylmethyl, furylmethyl, thienylethyl, pyrrolylalkyl, pyridylalkyl, isoxazollylalkyl, and imidazolylalkyl. In some cases, the alkylene group is a lower alkylene group (i.e., a C1-4 alkylene group).
As used herein, “carbocyclyl” means a non-aromatic cyclic ring or ring system containing only carbon atoms in the ring system backbone. When the carbocyclyl is a ring system, two or more rings may be joined together in a fused, bridged or spiro-connectedfashion. Carbocyclyls may have any degree of saturation provided that at least one ring in a ring system is not aromatic. Thus, carbocyclyls include cycloalkyls, cycloalkcnyls, and cycloalkynyls. The carbocyclyl group may have 3 to 20 carbon atoms, although the present definition also covers the occurrence of the term “carbocyclyl” where no numerical range is designated. The carbocyclyl group may also be a medium size carbocyclyl having 3 to 10 carbon atoms. The carbocyclyl group could also be a carbocyclyl having 3 to 6 carbon atoms. The carbocyclyl group may be designated as “C3-6 carbocyclyl” or similar designations. Examples of carbocyclyl rings include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cyclohexenyl, 2,3-dihydro-indene, bicycle[2.2.2]octanyl, adamantyl, and Spiro[4.4]nonanyl
A “(carbocyclyl)alkyl” is a carbocyclyl group connected, as a substituent, via an alkylene group, such as “C4-10 (carbocyclyl)alkyl” and the like, including but not limited to, cyclopropylmethyl, cyclobutylmethyl, cyclopropylethyl, cyclopropylbutyl, cyclobutylethyl, cyclopropylisopropyl, cyclopentylmethyl, cyclopentylethyl, cyclohexylmethyl, cyclohexylcthyl, cycloheptylmethyl, and the like. In some cases, the alkylene group is a lower alkylene group.
As used herein, “cycloalkyl” means a fully saturated carbocyclyl ring or ring system. Examples include cyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl.
As used herein, “cycloalkenyl” means a carbocyclyl ring or ring system having at least one double bond, wherein no ring in the ring system is aromatic. An example is cyclohexenyl.
As used herein, “heterocyclyl” means a non-aromatic cyclic ring or ring system containing at least one heteroatom in the ring backbone. Heterocyclyls may be joined together in a fused, bridged or spino-connected fashion. Heterocyclyls may have any degree of saturation provided that at least one ring in the ring system is not aromatic. The heteroatom(s) may be present in either a non-aromatic or aromatic ring in the ring system. The heterocyclyl group may have 3 to 20 ring members (i.e., the number of atoms making up the ring backbone, including carbon atoms and heteroatoms), although the present definition also covers the occurrence of the term “heterocyclyl” where no numerical range is designated. The heterocyclyl group may also be a medium size heterocyclyl having 3 to 10 ring members. The heterocyclyl group could also be a heterocyclyl having 3 to 6 ring members. The heterocyclyl group may be designated as “3-6 membered heterocyclyl” or similar designations. In preferred six membered monocyclic heterocyclyls, the heteroatom(s) are selected from one up to three of O (oxygen), N (nitrogen) or S (sulfur), and in preferred five membered monocyclic heterocyclyls, the heteroatom(s) are selected from one or two heteroatoms selected from O (oxygen), N (nitrogen), or S (sulfur). Examples of heterocyclyl rings include, but are not limited to, azepinyl, acridinyl, carbazolyl, cinnolinyl, dioxolanyl, imidazolinyl, imidazolidinyl, morpholinyl, oxiranyl, oxepanyl, thiepanyl, piperidinyl, piperazinyl, dioxopiperazinyl, pyrrolidinyl, pyrrolidonyl, pyrrolidionyl, 4-piperidonyl, pyrazolinyl, pyrazolidinyl, 1,3-dioxinyl, 1,3-dioxanyl, 1,4-dioxinyl, 1,4-dioxanyl, 1,3-oxathianyl, 1,4-oxathiinyl, 1,4-oxathianyl, 2H-1,2-oxazinyl, trioxanyl, hexahydro-1,3,5-triazinyl, 1,3-dioxolyl, 1,3-dioxolanyl, 1,3-dithiolyl, 1,3-dithiolanyl, isoxazolinyl, isoxazolidinyl, oxazolinyl, oxazolidinyl, oxazolidinonyl, thiazolinyl, thiazolidinyl, 1,3-oxathiolanyl, indolinyl, isoindolinyl, tetrahydrofuranyl, tetrahydropyranyl, tetrahydrothiophenyl, tetrahydrothiopyranyl, tetrahydro-1,4-thiazinyl, thiamorpholinyl, dihydrobenzofuranyl, benzimidazolidinyl, and tetrahydroquinoline.
A “(heterocyclyl)alkyl” is a heterocyclyl group connected, as a substituent, via an alkylene group. Examples include, but are not limited to, imidazolinylmethyl and indolinylethyl.
As used herein, “acyl” refers to —C(═O)R, wherein R is hydrogen, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C3-7 carbocyclyl, C6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein. Non-limiting examples include formyl, acetyl, propanoyl, benzoyl, and acryl.
An “O-carboxy” group refers to a “—OC(═O)R” group in which R is selected from hydrogen, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C3-7 carbocyclyl, C6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
A “C-carboxy” group refers to a “—C(═O)OR” group in which R is selected from hydrogen, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C3-7 carbocyclyl, C6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein. A non-limiting example includes carboxyl (i.e., —C(═O)OH).
A “cyano” group refers to a “—CN” group.
A “cyanato” group refers to an “—OCN” group.
An “isocyanato” group refers to a “—NCO” group.
A “thiocyanato” group refers to a “—SCN” group.
An “isothiocyanato” group refers to an “—NCS” group.
A “sulfinyl” group refers to an “—S(═O)R” group in which R is selected from hydrogen, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C3-7 carbocyclyl, C6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
A “sulfonyl” group refers to an “—SO2R” group in which R is selected from hydrogen, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C3-7 carbocyclyl, C6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
An “S-sulfonamido” group refers to a “—SO2NRARB” group in which RA and RB are each independently selected from hydrogen, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C3-7 carbocyclyl, C6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
An “N-sulfonamido” group refers to a “—N(RA)SO2RB” group in which RA and Rb are each independently selected from hydrogen, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C3-7 carbocyclyl, C6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
An “O-carbamyl” group refers to a “—OC(═O)NRARB” group in which RA and RB are each independently selected from hydrogen, C1-6 alkyl, C2-6 alkenyl, C2-6alkynyl, C3-7 carbocyclyl, C6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
An “N-carbamyl” group refers to an “—N(RA)C(═O)ORB” group in which RA and RB are each independently selected from hydrogen, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C3-7 carbocyclyl, C6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
An “O-thiocarbamyl” group refers to a “—OC(═S)NRARB” group in which RA and RB are each independently selected from hydrogen, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C3-7 carbocyclyl, C6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
A “urea” group refers to a “—N(RA)C(═O)NRARB” group in which RA and RB are each independently selected from hydrogen, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C3-7 carbocyclyl, C6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
An “N-thiocarbamyl” group refers to an “—N(RA)C(═S)ORB” group in which RA and RB are each independently selected from hydrogen, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C3-7 carbocyclyl, C6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
A “C-amido” group refers to a “—C(═O)NRARB” group in which RA and RB are each independently selected from hydrogen, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C3-7 carbocyclyl, C6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
An “N-amido” group refers to a “—N(RA)C(═O)RB” group in which RA and RB are each independently selected from hydrogen, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C3-7 carbocyclyl, C6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein.
An “amino” group refers to a “—NRARB” group in which RA and RB are each independently selected from hydrogen, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C3-7 carbocyclyl, C6-10 aryl, 5-10 membered heteroaryl, and 5-10 membered heterocyclyl, as defined herein. A non-limiting example includes free amino (i.e., —NH2).
An “aminoalkyl” group refers to an amino group connected via an alkylene group.
An “alkoxyalkyl” group refers to an alkoxy group connected via an alkylene group, such as a “C2-8 alkoxyalkyl” and the like.
As used herein, a substituted group is derived from the unsubstituted parent group in which there has been an exchange of one or more hydrogen atoms for another atom or group. Unless otherwise indicated, when a group is deemed to be “substituted,” it is meant that the group is substituted with one or more substitutents independently selected from C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 carbocyclyl (optionally substituted with halo, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 haloalkyl, and C1-C6 haloalkoxy), C3-C7-carbocyclyl-C1-C6-alkyl (optionally substituted with halo, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 haloalkyl, and C1-C6 haloalkoxy), 5-10 membered heterocyclyl (optionally substituted with halo, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 haloalkyl, and C1-C6 haloalkoxy), 5-10 membered heterocyclyl-C1-C6-alkyl (optionally substituted with halo, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 haloalkyl, and C1-C6 haloalkoxy), aryl (optionally substituted with halo, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 haloalkyl, and C1-C6 haloalkoxy), aryl(C1-C6)alkyl (optionally substituted with halo, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 haloalkyl, and C1-C6 haloalkoxy), 5-10 membered heteroaryl (optionally substituted with halo, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 haloalkyl, and C1-C6 haloalkoxy), 5-10 membered heteroaryl(C1-C6)alkyl (optionally substituted with halo, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 haloalkyl, and C1-C6 haloalkoxy), halo, cyano, hydroxy, C1-C6 alkoxy, C1-C6 alkoxy(C1-C6)alkyl (i.e., ether), aryloxy, sulfhydryl (mercapto), halo(C1-C6)alkyl (e.g., —CF3), halo(C1-C6)alkoxy (e.g., —OCF3), C1-C6 alkylthio, arylthio, amino, amino(C1-C6)alkyl, nitro, O-carbamyl, N-carbamyl, O-thiocarbamyl, N-thiocarbamyl, C-amido, N-amido, S-sulfonamido, N-sulfonamido, C-carboxy, O-carboxy, acyl, cyanato, isocyanato, thiocyanato, isothiocyanato, sulfinyl, sulfonyl, and oxo (═O). Wherever a group is described as “optionally substituted” that group can be substituted with the above substituents.
It is to be understood that certain radical naming conventions can include either a mono-radical or a di-radical, depending on the context. For example, where a substituent requires two points of attachment to the rest of the molecule, it is understood that the substituent is a di-radical. For example, a substituent identified as alkyl that requires two points of attachment includes di-radicals such as —CH2—, —CH2CH2—, —CH2CH(CH3)CH2—, and the like. Other radical naming conventions clearly indicate that the radical is a di-radical such as “alkylene” or “alkenylene.”
Wherever a substituent is depicted as a di-radical (i.e., has two points of attachment to the rest of the molecule), it is to be understood that the substituent can be attached in any directional configuration unless otherwise indicated. Thus, for example, a substituent depicted as -AE- or
Figure US12115228-20241015-C00005
    •  includes the substituent being oriented such that the A is attached at the leftmost attachment point of the molecule as well as the case in which A is attached at the rightmost attachment point of the molecule.
“Subject” as used herein, means a human or a non-human mammal, e.g., a dog, a cat, a mouse, a rat, a cow, a sheep, a pig, a goat, a non-human primate or a bird, e.g., a chicken, as well as any other vertebrate or invertebrate.
Conjugation Methods, Spacers and Linkers Involved
Some embodiments provide a method of conjugating of a targeting molecule through a spacer or a multifunctional linker. In some embodiments, the spacer or multifunctional linker may include a 2- to 5-atom bridge. In some embodiments, the method includes a single-step or sequential conjugation approach. In some embodiments, the drug-conjugates include a spacer or a multifunctional linker. In some embodiments, the spacer or multifunctional linker may include a noncleavable or cleavable unit such as peptides.
Utilities and Applications
Some embodiments provide a method of treating a patient in need thereof comprising administering an active agent-conjugate as disclosed and described herein to said patient. In some embodiments, the patient may have cancer, immune diseases or diabetes.
Some embodiments provide a method of diagnosis or imaging comprising administering an active agent-conjugate as disclosed and described herein to an individual.
Certain Structures
Some embodiments provide an active agent-conjugate having the structure of Formula I
Figure US12115228-20241015-C00006
    • or a pharmaceutically acceptable salt thereof,
    • wherein:
    • A may be a targeting molecule;
    • B is an auxiliary moiety that optionally is a second targeting molecule, or B is null;
    • L1 includes a 2- to 5-carbon bridge and at least one sulfur atom;
    • each D is independently selected, where each D includes an active agent;
    • each L2 is independently a linker, wherein at least one L2 links to L1; and
    • n is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.
In some embodiments, A may be a monoclonal antibody (mAB).
In some embodiments, A may be an antibody fragment, surrogate, or variant.
In some embodiments, A may be a protein ligand.
In some embodiments, A may be a protein scaffold.
In some embodiments, A may be a peptide.
In some embodiments, A may be a small molecule ligand.
In some embodiments, B may be a hydrophilic polymer. In some embodiments, the hydrophilic polymer may polyethylene glycol (PEG), and the like. In some embodiments, B may be a biodegradable polymer. In some embodiments, the biodegradable polymer may be unstructured proteins polyamino acids, polypeptides polysaccharides and combinations thereof.
In some embodiments, B may be a monoclonal antibody (mAB).
In some embodiments, B may be an antibody fragment, surrogate, or variant.
In some embodiments, B may be a protein ligand.
In some embodiments, B may be a protein scaffold.
In some embodiments, B may be a peptide.
In some embodiments, B may be RNA or DNA.
In some embodiments, B may be a RNA or DNA fragment.
In some embodiments, B may be a small molecule ligand.
In some embodiments, D may be a biologically active compound.
In some embodiments, D may be a drug.
In some embodiments, D may be a chemotherapy drug.
In some embodiments, D may be a natural product.
In some embodiments, D may be an immune modulator.
In some embodiments, D may be a tubulin-binder.
In some embodiments, D may be a DNA-alkylating agent.
In some embodiments, D may be an HSP90 inhibitor.
In some embodiments, D may be a DNA topoisomerase inhibitor.
In some embodiments, D may be an anti-epigenetic agent.
In some embodiments, D may be an HDAC inhibitor.
In some embodiments, D may be an anti-metabolism agent.
In some embodiments, D may be a proteasome inhibitor.
In some embodiments, D may be a peptide.
In some embodiments, D may be a peptidomimetic.
In some embodiments, D may be an siRNA.
In some embodiments, D may be an antisense DNA.
In some embodiments, D may be epothilone A, epothilone B, or paclitaxel.
In some embodiments, L2 may include a spacer or a multifunctional linker. In some embodiments, L2 may include a spacer and a multifunctional linker. In some embodiments, L2 may include a multifunctional linker. In some embodiments, each L2 may be a linker, wherein the linker may be cleavable or non-cleavable under biological conditions. In some embodiments, the linker may be cleavable by an enzyme. In some embodiments, L2 may include Linker.
In some embodiments, L1 includes a 2- to 5-carbon bridge and at least one sulfur atom. In some embodiments, L1 includes a 2- to 5-carbon bridge and at least two sulfur atoms. In some embodiments, L1 includes a 2- to 5-carbon bridge and a spacer. In some embodiments, L1 includes a 2- to 5-carbon bridge, at least two sulfur atoms and a spacer. In some embodiments, L1 may include one or more sulfurs. In some embodiments, the L1 may include two or more sulfurs. In some embodiments, the L1 may include exactly two sulfurs. In some embodiments, may include a 4-carbon bridge and/or a spacer. In some embodiments, L1 include a 4-carbon bridge or a spacer. In some embodiments, L1 may include a 4-carbon bridge and a spacer. In some embodiments, L1 includes a 4-carbon bridge and at least two sulfur atoms. In some embodiments, the spacer connects to the mAB by a sulfide bond. In some embodiments, the spacer connects to the mAB through a thioether.
In some embodiments, A comprises at least one modified L-Alanine residue. In some embodiments, A comprises at least two modified L-Alanine residues. In some embodiments, L1 comprises at least one sulfur. In some embodiments, L1 comprises at least two sulfurs. In some embodiments, A comprises at least one modified L-Alanine residue that is connected to at least one sulfur of L1. In some embodiments, at least one modified L-Alanine residue is from an L-Cysteine residue of a peptide before conjugation. In some embodiments, at least one sulfur of L1 is from an L-Cysteine of a peptide before conjugation. In some embodiments, A and L1 together comprise at least one thioether.
In some embodiments, Linker may be a peptide.
In some embodiments, Linker may include an oligosaccharide. For example, Linker may include chitosan. In some embodiments, L2 may include Linker and —(CH2)n— where n is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. In some embodiments, L2 may include Linker and —(CH2CH2O)n— where n is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.
In some embodiments, Linker may include —(CH2)n— where n is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.
In some embodiments, Linker may include —(CH2CH2O)n— where n is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.
In some embodiments, Linker may include Val-Cit-PAB, Val-Ala-PAB, Val-Lys(Ac)-PAB, Phe-Lys-PAB, Phe-Lys(Ac)-PAB, D-Val-Leu-Lys, Gly-Gly-Arg, Ala-Ala-Asn-PAB, Ala-PAB, PAB, or the like.
In some embodiments, Linker may include any combination of peptide, oligosaccharide, —(CH2)n—, —(CH2CH2O)n—, Val-Cit-PAB, Val-Ala-PAB, Val-Lys(Ac)-PAB, Phe-Lys-PAB, Phe-Lys(Ac)-PAB, D-Val-Leu-Lys, Gly-Gly-Arg, Ala-Ala-Asn-PAB, Ala-PAB, PAB, and the like.
In some embodiments, the spacer may include a peptide.
In some embodiments, the spacer may include an oligosaccharide. For example, the spacer may include chitosan.
In some embodiments, the spacer may include —(CH2)n— where n is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. In some embodiments, L1 may include a component including a 4-carbon bridge and —(CH2)n— where n is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.
In some embodiments, the spacer may include —(CH2CH2O)n— where n is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. In some embodiments, L1 may include a component including a 4-carbon bridge and —(CH2CH2O)n— where n is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.
In some embodiments, the spacer may include Val-Cit-PAB, Val-Ala-PAB, Val-Lys(Ac)-PAB, Phe-Lys-PAB, Phe-Lys(Ac)-PAB, D-Val-Leu-Lys, Gly-Gly-Arg, Ala-Ala-Asn-PAB, Ala-PAB, PAB, or the like.
In some embodiments, the spacer may be any combination of peptide, oligosaccharide, —(CH2)n—, —(CH2CH2O)n—, Val-Cit-PAB, Val-Ala-PAB, Val-Lys(Ac)-PAB, Phe-Lys-PAB, Phe-Lys(Ac)-PAB, D-Val-Leu-Lys, Gly-Gly-Arg, Ala-Ala-Asn-PAB, Ala-PAB, PAB, and the like.
In some embodiments, L1 may include a 4-carbon bridge. In some embodiments, L1 may include a 4-carbon bridge through a fused aromatic ring. In some embodiments, L1 may include a spacer and component including a 4-carbon bridge through a fused aromatic ring. In some embodiments, L1 may include, but is not limited
Figure US12115228-20241015-C00007
    •  and the like.
In some embodiments, the agent-conjugate having the structure of Formula I has the structure of Formula Ia:
Figure US12115228-20241015-C00008
    • or a pharmaceutically acceptable salt thereof,
    • wherein the S-linked portion of A comprises a modified L-Alanine residue, wherein
Figure US12115228-20241015-C00009
    •  may connect to a sulfur of a reduced disulfide bond through a bridge containing 2 to 5 atoms. For example, the structure indicated by
Figure US12115228-20241015-C00010
    •  may include a fragment selected from the group consisting of:
Figure US12115228-20241015-C00011
In some embodiments, the S-linked (sulfur-linked) portion of A comprises a modified L-Alanine residue. In some embodiments, the S-linked (sulfur-linked) portion of A comprises a modified L-Alanine residue that is connected to the sulfur of L1. In some embodiments, the S-linked (sulfur-linked) portion of A comprises a modified L-Alanine residue wherein the modified L-Alanine component of A is from an L-Cysteine residue of a peptide before conjugation. In some embodiments, the sulfur of L1 is from an L-Cysteine of a peptide before conjugation. In some embodiments, A and L1 comprise a thioether. In some embodiments, the compound having the structure of Formula I a may be formed from a peptide that includes at least one L-Cysteine where the L-Cysteine provides a modified L-Alanine of A and a sulfur of
Figure US12115228-20241015-C00012
    •  In some embodiments, the compound having the structure of Formula Ia may be formed from a peptide that includes at least two L-Cysteine where the two L-Cysteine provide two modified L-Alanines of A and two sulfur of
Figure US12115228-20241015-C00013
In some embodiments, the agent-conjugate having the structure of Formula I has the structure of Formula Ib:
Figure US12115228-20241015-C00014
    • or a pharmaceutically acceptable salt thereof, wherein the A-component may be a targeting moiety, the E-component may be a heteroaryl or heterocyclyl, L3 may be an optionally substituted C1-C6 alkyl, or L3 may be null, when L3 is null the sulfur is directly connected to the E-component; and L4 may be an optionally substituted C1-C6 alkyl, or L4 may be null, when L4 is null the sulfur is directly connected to the E-component. In some embodiments, L3 may be —(CH2)—; and L4 may be —(CH2)—. In some embodiments, L3 may be null; and L4 may be null.
In some embodiments of the structure of Formula Ib, the structural component
Figure US12115228-20241015-C00015
    • may be:
Figure US12115228-20241015-C00016
In some embodiments, the E-component includes a fragment selected from the group consisting of:
Figure US12115228-20241015-C00017
In some embodiments, the active agent may be selected from the group consisting of tubulin binders, DNA alkylators, DNA intercalator, enzyme inhibitors, immune modulators, peptides, and nucleotides.
In some embodiments, at least one L2 includes —(CH2)n— where n is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. In some embodiments, at least one L2 includes —(CH2CH2O)n— where n is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. In some embodiments, at least one L2 includes Val-Cit-PAB, Val-Ala-PAB, Val-Lys(Ac)-PAB, Phe-Lys-PAB, Phe-Lys(Ac)-PAB, D-Val-Leu-Lys, Gly-Gly-Arg, Ala-Ala-Asn-PAB, Ala-PAB, or PAB. In some embodiments, at least one L2 includes a peptide, an oligosaccharide, —(CH2)n—, —(CH2CH2O)n—, Val-Cit-PAB, Val-Ala-PAB, Val-Lys(Ac)-PAB, Phe-Lys-PAB, Phe-Lys(Ac)-PAB, D-Val-Leu-Lys, Gly-Gly-Arg, Ala-Ala-Asn-PAB, Ala-PAB, or PAB. In some embodiments, at least one L2 may comprise, consist of, or consist essentially of:
Figure US12115228-20241015-C00018
Figure US12115228-20241015-C00019
Figure US12115228-20241015-C00020
Figure US12115228-20241015-C00021
Figure US12115228-20241015-C00022
Figure US12115228-20241015-C00023
Figure US12115228-20241015-C00024
In some embodiments, the agent-conjugate having the structure of Formula I has the structure of
Figure US12115228-20241015-C00025
In some embodiments, the agent-conjugate having the structure of Formula I has the structure of
Figure US12115228-20241015-C00026
In some embodiments, the agent-conjugate having the structure of Formula I has the structure of
Figure US12115228-20241015-C00027
In some embodiments, the agent-conjugate having the structure of Formula I has the structure of
Figure US12115228-20241015-C00028
In some embodiments, the agent-conjugate having the structure of Formula I has the structure of
Figure US12115228-20241015-C00029
In some embodiments, the agent-conjugate having the structure of Formula I has the structure of
Figure US12115228-20241015-C00030
In some embodiments, the agent-conjugate having the structure of Formula I has the structure of
Figure US12115228-20241015-C00031
In some embodiments, the agent-conjugate having the structure of Formula I has the structure of
Figure US12115228-20241015-C00032
In some embodiments, the agent-conjugate having the structure of Formula I has the structure of
Figure US12115228-20241015-C00033
In some embodiments, the agent-conjugates may include one or more components selected from the group consisting of an amino acid, an amino acid residue, an amino acid analog, and a modified amino acid.
As used herein, the term “targeting moiety” refers to a structure that binds or associates with a biological moiety or fragment thereof.
In some embodiments, the targeting moiety may be an antibody. In some embodiments, the targeting moiety may be a monoclonal antibody (mAB). In some embodiments, the targeting moiety may be an antibody fragment, surrogate, or variant. In some embodiments, the targeting moiety may be a protein ligand. In some embodiments, the targeting moiety may be a protein scaffold. In some embodiments, the targeting moiety may be a peptide. In some embodiments, the targeting moiety may be RNA or DNA. In some embodiments, the targeting moiety may be a RNA or DNA fragment. In some embodiments, the targeting moiety may be a small molecule ligand.
In some embodiments, the targeting moiety may be an antibody fragment described in Janthur et al., “Drug Conjugates Such as Antibody Drug Conjugates (ADCs), Immunotoxins and Immunoliposomes Challenge Daily Clinical Practice,” Int. J. Mol. Sci. 2012, 13, 16020-16045, the disclosure of which is incorporated herein by reference in its entirety. In some embodiments, the targeting moiety may be an antibody fragment described in Trail, PA, “Antibody Drug Conjugates as Cancer Therapeutics,” Antibodies 2013, 2, 113-129, the disclosure of which is incorporated herein by reference in its entirety.
In some embodiments, the targeting moiety may be HuM195-Ac-225, HuM195-Bi-213, Anyara (naptumomab estafenatox; ABR-217620), AS1409, Zevalin (ibritumomab tiuxetan), BIIB015, BT-062, Neuradiab, CDX-1307, CR011-vcMMAE, Trastuzumab-DM1 (R3502), Bexxar (tositumomab), IMGN242, IMGN388, 1MGN901, 131I-labetuzumab, IMMU-102 (90Y-epratuzumab), IMMU-107 (90Y-clivatuzumab tetraxetan), MDX-1203, CAT-8015, EMD 273063 (hu14.18-1L2), Tucotuzumab celmoleukin (EMD 273066; huKS-IL2), 188Re-PTI-6D2, Cotara, L19-IL2, Teleukin (F16-IL2), Tenarad (F16-131I), L19-131I, L19-TNF, PSMA-ADC, DI-Leu16-IL2, SAR3419, SGN-35, or CMC544. In some embodiments, the targeting moiety may comprise, consist of, or consist essentially of the antibody portion of HuM195-Ac-225, HuM195-Bi-213, Anyara (naptumomab estafenatox; ABR-217620), AS1409, Zevalin (ibritumomab tiuxetan), BIIB015, BT-062, Neuradiab, CDX-1307, CR011-vcMMAE, Trastuzumab-DM1 (R3502), Bexxar (tositumomab), IMGN242, IMGN388, IMGN901, 131I-labetuzumab, IMMU-102 (90Y-epratuzumab), IMMU-107 (90Y-clivatuzumab tetraxetan), MDX-1203, CAT-8015, EMD 273063 (hu14.18-1L2), Tucotuzumab celmoleukin (EMD 273066; huKS-1L2), 188Re-PTI-6D2, Cotara, L19-1L2, Teleukin (F16-1L2), Tenarad (F16-131I), L19-131I, L19-TNF, PSMA-ADC, DI-Leu16-IL2, SAR3419, SGN-35, or CMC544.
In some embodiments, the targeting moiety may be Brentuximab vedotin, Trastuzumab emtansine, Inotuzumab ozogamicin, Lorvotuzumab mertansine, Glembatumumab vedotin, SAR3419, Moxetumomab pasudotox, Moxetumomab pasudotox, AGS-16M8F, AGS-16M8F, BIIB-015, BT-062, IMGN-388, or IMGN-388.
In some embodiments, the targeting moiety may comprise, consist of, or consist essentially of the antibody portion of Brentuximab vedotin, Trastuzumab emtansine, Inotuzumab ozogamicin, Lorvotuzumab mertansine, Glembatumumab vedotin, SAR3419, Moxetumomab pasudotox, Moxetumomab pasudotox, AGS-16M8F, AGS-16M8F, BIIB-015, BT-062, IMGN-388, or IMGN-388.
In some embodiments, the targeting moiety may comprise, consist of, or consist essentially of Brentuximab, Inotuzumab, Gemtuzumab, Milatuzumab, Trastuzumab, Glembatumomab, Lorvotuzumab, or Lab estuzumab.
As used herein, the term “peptide” refers to a structure including one or more components each individually selected from the group consisting of an amino acid, an amino acid residue, an amino acid analog, and a modified amino acid. The components are typically joined to each other through an amide bond.
In some embodiments, the peptide, such as the antibody, is PEGylated. PEGylation can provide, for example, increased stability and/or efficacy of the polypeptide. Methods for PEGylation known in the art can be used in the methods and compositions provided herein. Such methods include, but are not limited to, those provided in “Comparative Binding of Disulfide-Bridged PEG-Fabs” Khalili et al., Bioconjugate Chemistry (2012), 23(11), 2262-2277; “Site-Specific PEGylation at Histidine Tags” Cong et al., Bioconjugate Chemistry (2012), 23(2), 248-263; “Disulfide bridge based PEGylation of proteins” Brocchini et al., Advanced Drug Delivery Reviews (2008), 60(1), 3-12; “Site-Specific PEGylation of Protein Disulfide Bonds Using a Three-Carbon Bridge” Balan et al., Bioconjugate Chemistry (2007), 18(1), 61-76; “Site-specific PEGylation of native disulfide bonds in therapeutic proteins” Shaunak et al., Nature Chemical Biology (2006), 2(6), 312-313; “PEG derivative conjugated proteins and peptides for use in pharmaceuticals” Godwin et al., WO 2010/100430. All of the aforementioned PEGylation references are incorporated by references in their entireties.
As used herein, the term “amino acid” includes naturally occurring amino acids, a molecule having a nitrogen available for forming an amide bond and a carboxylic acid, a molecule of the general formula NH2—CHR—COOH or the residue within a peptide bearing the parent amino acid, where “R” is one of a number of different side chains. “R” can be a substituent found in naturally occurring amino acids. “R” can also be a substituent referring to one that is not of the naturally occurring amino acids.
As used herein, the term “amino acid residue” refers to the portion of the amino acid which remains after losing a water molecule when it is joined to another amino acid.
As used herein, the term “amino acid analog” refers to a structural derivative of an amino acid parent compound that often differs from it by a single element.
As used herein, the term “modified amino acid” refers to an amino acid bearing an “R” substituent that does not correspond to one of the twenty genetically coded amino acids.
As used herein, the abbreviations for the genetically encoded L-enantiomeric amino acids are conventional and are as follows: The D-amino acids are designated by lower case, e.g. D-proline=p, etc.
TABLE 1
One-Letter Common
Amino Acids Symbol Abbreviation
Alanine A Ala
Arginine R Arg
Asparagine N Asn
Aspartic acid D Asp
Cysteine C Cys
Glutamine Q Gln
Glutamic acid E Glu
Glycine G Gly
Histidine H His
Isoleucine I Ile
Leucine L Leu
Lysine K Lys
Phenylalanine F Phe
Proline P Pro
Serine S Ser
Threonine T Thr
Tryptophan W Trp
Tyrosine Y Tyr
Valine V Val
Certain amino acid residues in the active agent-conjugate can be replaced with other amino acid residues without significantly deleteriously affecting, and in many cases even enhancing, the activity of the peptides. Thus, also contemplated by the preferred embodiments are altered or mutated forms of the active agent-conjugate wherein at least one defined amino acid residue in the structure is substituted with another amino acid residue or derivative and/or analog thereof. It will be recognized that in preferred embodiments, the amino acid substitutions are conservative, i.e., the replacing amino acid residue has physical and chemical properties that are similar to the amino acid residue being replaced.
For purposes of determining conservative amino acid substitutions, the amino acids can be conveniently classified into two main categories—hydrophilic and hydrophobic—depending primarily on the physical-chemical characteristics of the amino acid side chain. These two main categories can be further classified into subcategories that more distinctly define the characteristics of the amino acid side chains. For example, the class of hydrophilic amino acids can be further subdivided into acidic, basic and polar amino acids. The class of hydrophobic amino acids can be further subdivided into nonpolar and aromatic amino acids. The definitions of the various categories of amino acids are as follows:
The term “hydrophilic amino acid” refers to an amino acid exhibiting a hydrophobicity of less than zero according to the normalized consensus hydrophobicity scale of Eisenberg et al., 1984, J. Mol. Biol. 179:125-142. Genetically encoded hydrophilic amino acids include Thr (T), Ser (S), His (H), Glu (E), Asn (N), Gln (Q), Asp (D), Lys (K) and Arg (R).
The term “hydrophobic amino acid” refers to an amino acid exhibiting a hydrophobicity of greater than zero according to the normalized consensus hydrophobicity scale of Eisenberg, 1984, J. Mol. Biol. 179:1.25-142. Genetically encoded hydrophobic amino acids include Pro (P), Ile (I), Phe (F), Val (V), Leu (L), Trp (W), Met (M), Ala (A), Gly (G) and Tyr (Y).
The term “acidic amino acid” refers to a hydrophilic amino acid having a side chain pK value of less than 7. Acidic amino acids typically have negatively charged side chains at physiological pH due to loss of a hydrogen ion. Genetically encoded acidic amino acids include Glu (E) and Asp (D).
The term “basic amino acid” refers to a hydrophilic amino acid having a side chain pK value of greater than 7. Basic amino acids typically have positively charged side chains at physiological pH due to association with hydronium ion. Genetically encoded basic amino acids include His (H), Arg (R) and Lys (K).
The term “polar amino acid” refers to a hydrophilic amino acid having a side chain that is uncharged at physiological pH, but which has at least one bond in which the pair of electrons shared in common by two atoms is held more closely by one of the atoms. Genetically encoded polar amino acids include Asn (N), Gln (Q) Ser (S) and Thr (T).
The term “nonpolar amino acid” refers to a hydrophobic amino acid having a side chain that is uncharged at physiological pH and which has bonds in which the pair of electrons shared in common by two atoms is generally held equally by each of the two atoms (i.e., the side chain is not polar). Genetically encoded nonpolar amino acids include Leu (L), Val (V), Ile (I), Met (M), Gly (G) and Ala (A).
The term “aromatic amino acid” refers to a hydrophobic amino acid with a side chain having at least one aromatic or heteroaromatic ring. In some embodiments, the aromatic or heteroaromatic ring may contain one or more substituents such as OH, SH, CN, F, Cl, Br, I, NO2, NO, NH2, NHR, NRR, —C(O)R, —C(O)OH, —C(O)OR, —C(O)NH2, —C(O)NHR, —C(O)NRR and the like where each R is independently (C1-C6) alkyl, substituted (C1-C6) alkyl, (C1-C6) alkenyl, substituted (C1-C6) alkenyl, (C1-C6) alkynyl, substituted (C1-C6) alkynyl, (C5-C20) aryl, substituted (C5-C20) aryl, (C6-C26) alkaryl, substituted (C6-C26) alkaryl, 5-20 membered heteroaryl, substituted 5-20 membered heteroaryl, 6-26 membered alkheteroaryl or substituted 6-26 membered alkheteroaryl. Genetically encoded aromatic amino acids include Phe (F), Tyr (Y) and Trp (W).
The term “aliphatic amino acid” refers to a hydrophobic amino acid having an aliphatic hydrocarbon side chain. Genetically encoded aliphatic amino acids include Ala (A), Val (V), Leu (L) and Ile (I).
The amino acid residue Cys (C) is unusual in that it can form disulfide bridges with other Cys (C) residues or other sulfanyl-containing amino acids. The ability of Cys (C) residues (and other amino acids with —SH containing side chains) to exist in a peptide in either the reduced free —SH or oxidized disulfide-bridged form affects whether Cys (C) residues contribute net hydrophobic or hydrophilic character to a peptide. While Cys (C) exhibits a hydrophobicity of 0.29 according to the normalized consensus scale of Eisenberg (Eisenberg, 1984, supra), it is to be understood that for purposes of the preferred embodiments Cys (C) is categorized as a polar hydrophilic amino acid, notwithstanding the general classifications defined above.
As will be appreciated by those of skill in the art, the above-defined categories are not mutually exclusive. Thus, amino acids having side chains exhibiting two or more physical-chemical properties can be included in multiple categories. For example, amino acid side chains having aromatic moieties that are further substituted with polar substituents, such as Tyr (Y), may exhibit both aromatic hydrophobic properties and polar or hydrophilic properties, and can therefore be included in both the aromatic and polar categories. The appropriate categorization of any amino acid will be apparent to those of skill in the art, especially in light of the detailed disclosure provided herein.
While the above-defined categories have been exemplified in terms of the genetically encoded amino acids, the amino acid substitutions need not be, and in certain embodiments preferably are not, restricted to the genetically encoded amino acids. In some embodiments, the active agent-conjugate may contain genetically non-encoded amino acids. Thus, in addition to the naturally occurring genetically encoded amino acids, amino acid residues in the active agent-conjugate may be substituted with naturally occurring non-encoded amino acids and synthetic amino acids.
Certain commonly encountered amino acids which provide useful substitutions for the active agent-conjugates include, but are not limited to, β-alanine (β-Ala) and other omega-amino acids such as 3-aminopropionic acid, 2,3-diaminopropionic acid (Dpr), 4-aminobutyric acid and so forth; α-aminoisobutyric acid (Aib); ε-aminohexanoic acid (Aha); δ-aminovaleric acid (Ava); N-methylglycine or sarcosine (MeGly); ornithine (Orn); citrullinc (Cit); t-butylalaninc (t-BuA); t-butylglycinc (t-BuG); N-methylisoleucine (Melle); phenylglycine (Phg); cyclohexylalanine (Cha); norleucine (Nle); naphthylalanine (Nal); 4-phenylphenylalanine, 4-chlorophenylalanine (Phe(4-C1)); 2-fluorophenylalanine (Phe(2-F)); 3-fluorophenylalanine (Phe(3-F)); 4-fluorophenylalanine (Phe(4-F)); penicillamine (Pen); 1,2,3,4-tetrahydroisoquinoline-3-carboxylic acid (Tic); 13-2-thienylalanine (Thi); methionine sulfoxidc (MSO); homoarginine (hArg); N-acetyl lysine (AcLys); 2,4-diaminobutyric acid (Dbu); 2,3-diaminobutyric acid (Dab); p-aminophenylalanine (Phe (pNH2)); N-methyl valine (MeVal); homocysteine (hCys), homophenylalanine (hPhe) and homoserine (hSer); hydroxyproline (Hyp), homoproline (hPro), N-methylated amino acids and peptoids (N-substituted glycines).
Other amino acid residues not specifically mentioned herein can be readily categorized based on their observed physical and chemical properties in light of the definitions provided herein.
The classifications of the genetically encoded and common non-encoded amino acids according to the categories defined above are summarized in Table 2, below. It is to be understood that Table 2 is for illustrative purposes only and does not purport to be an exhaustive list of amino acid residues and derivatives that can be used to substitute the active agent-conjugate described herein.
TABLE 2
CLASSIFICATIONS OF COMMONLY
ENCOUNTERED AMINO ACIDS
Genetically Non-Genetically
Classification Encoded Encoded
Hydrophobic
Aromatic F, Y, W Phg, Nal, Thi, Tic, Phe (4-Cl),
Phe (2-F), Phe (3-F), Phe (4-F), hPhe
Nonpolar L, V, I, M, G, t-BuA, t-BuG, MeIle, Nle, MeVal, Cha,
A, P McGly, Aib
Aliphatic A, V, L, I b-Ala, Dpr, Aib, Aha, MeGly, t-BuA,
t-BuG, MeIle, Cha, Nle, MeVal
Hydrophilic
Acidic D, E
Basic H, K, R Dpr, Orn, hArg, Phe(p-NH2), Dbu, Dab
Polar C, Q, N, S. T Cit, AcLys, MSO, bAla, hSer
Helix-Breaking P, G D-Pro and other D-amino acids (in
L-peptides)
Other amino acid residues not specifically mentioned herein can be readily categorized based on their observed physical and chemical properties in light of the definitions provided herein.
While in most instances, the amino acids of the active agent-conjugate will be substituted with L-enantiomeric amino acids, the substitutions are not limited to L-enantiomeric amino acids. In some embodiments, the peptides may advantageously be composed of at least one D-enantiomeric amino acid. Peptides containing such D-amino acids are thought to be more stable to degradation in the oral cavity, gut or serum than are peptides composed exclusively of L-amino acids.
Conjugation Methods
One-Step Conjugation
Figure US12115228-20241015-C00034
Two-Step Conjugation, Drug Preloaded
Figure US12115228-20241015-C00035
Two-Step Conjugation, Drug Added at the Last Step
Figure US12115228-20241015-C00036
Examples of One-Step Conjugation
General Procedure 1
Figure US12115228-20241015-C00037
Examples include, but are not limited:
Figure US12115228-20241015-C00038
Additional compounds that can be made according to the General procedures include, but are not limited to, the following:
Figure US12115228-20241015-C00039
Figure US12115228-20241015-C00040
Figure US12115228-20241015-C00041
Examples include, but are not limited, the following:
Figure US12115228-20241015-C00042
Two-step conjugation, drugs preloaded
Approach:
Figure US12115228-20241015-C00043
Examples Include, but are not Limited to, the Following:
Figure US12115228-20241015-C00044
Conjugation Method
Figure US12115228-20241015-C00045
Examples Include, but are not Limited to the Following:
Figure US12115228-20241015-C00046
Conjugation Method
Figure US12115228-20241015-C00047
Examples Include, but are not Limited to, the Following:
Figure US12115228-20241015-C00048
EXAMPLES
Antibody-Drug Conjugation
General conjugation procedure—To a target antibody, 0.5-50 mgs/mL, in a certain buffet at pH 5.0-9.0, such as PBS, was added 0.5-100 eq of reducing agent such as TCEP and DTT. The reduction was performed at 0-40° C. for 0.5-40 hours with gentle stirring or shaking, and then the reducing agent was removed by column or ultrafiltration. To the partially reduced antibody, 0.5-50 mgs/mL, in a certain buffet at pH 5.0-9.0, such as PBS, with 0-30% of organic co-solvent such as DMA, was added 0.5-10 eq of the activated drug-linker bearing a dibromo or dichloro-functionality. The reaction was conducted at 0-40° C. for 0.5-40 hours with gentle stirring or shaking, monitored by HIC-HPLC. The resultant crude ADC product underwent necessary down-stream steps of desalt, buffet changes/formulation, and optionally, purification, using the state-of-art procedures. The final ADC product was characterized by HIC-HPLC, SEC, RP-HPLC, and optionally LC-MS. The average DAR was calculated by UV absorption and/or MS spectroscopy.
General Synthetic Procedures
General Procedure A—HATU Mediated amide Bond Formation
To an acid (1.1 eq with respect to amine) in anhydrous DMF was added HATU (1 eq with respect to acid) and DIEA (2 eq with respect to acid) and the mixture was stirred at room temperature for 1 minute. The mixture was then added to a solution of amine in DMF and the reaction mixture was stirred at room temperature till the completion of the reaction (monitored by LC/MS). The solvent was removed under reduced pressure and the residue was optionally purified by reverse phase HPLC to give final pure product.
General Procedure B—DIC/HOAt Mediated amide Bond Formation
To a stirred solution of carboxylic acid (1.1 eq), amine and HOAt (1.1 eq) in anhydrous DMF was added DIC (1.1 eq) and the reaction mixture was stirred at room temperature. Upon completion (monitored by LC/MS), the solvent was removed under reduced pressure and the residue was optionally purified by reverse phase HPLC to give final pure product.
General Procedure C—Removal of Acid Sensitive Protecting Groups (Boc, THP, t-Bu) Using HCl/dioxane
The acid sensitive protecting groups containing compound was dissolved in 4N HCl/dioxane and the mixture was stirred at room temperature for 2 h. The solution was then concentrated under reduced pressure and the residue was washed twice with cold ether. Purification was carried out on reverse phase HPLC if necessary.
General Procedure D—Removal of Fmoc Group
The Fmoc containing compound was dissolved in 2-5% piperidine in DMF. The mixture was stirred at room temperature for 1 h. The solvents were removed under reduced pressure. Purification was carried out on reverse phase HPLC if necessary.
General Procedure E—Reductive Alkylation
An amine was dissolved in DMF and aldehyde (5 eq) was added, followed by addition of sodium cyanoborohydridc (5 eq). HOAc was added to adjust the pH of the reaction mixture to 4-5. The mixture was stirred at room temperature till completion (1-4 h, monitored by HPLC). Purification was carried out on reverse phase HPLC if necessary.
General Procedure F—Saponification—Removal of Me/Et from esters
To a stirred solution of an ester in MeOH was added 1M aq. solution of LiOH till pH of the mixture was about 13-14 and the reaction mixture was stirred at room temperature till completion (˜16 h, monitored by HPLC). Citric acid (˜10% aq) was added to neutralize the reaction and the solvents were removed under reduced pressure. The crude product was optionally purified by RP-HPLC or used directly in the next step.
General Procedure G—Activation of a hydroxyl/phenol Group with Bis(p-nitrophenyl)carbonate
To a stirred solution of an alcohol/phenol in THF/DMF (2/1) was added bis(p-nitrophenyl) carbonate (3-5 eq), followed by DIEA (2-4 eq) and the reaction mixture was stirred at room temperature until most of the starting material was consumed. The progress of the reaction was monitored by LC/MS. The crude product was optionally purified by flash column chromatography or by precipitation and washing.
General Procedure H—Reaction of an amine with a cyclic anhydride (glutaric anhydride or succinic anhydride)
An amine containing compound was dissolved in DMF. Glutaric anhydride (3 eq) was added, followed by addition of DTEA (4 eq). The reaction mixture was stirred at room temperature until most of the starting material was consumed. The progress of the reaction was monitored by LC/MS. The crude product was purified by RP-HPLC to yield the pure carboxylic acid.
General Procedure I—Formation of carbamate with p-nitrophenyl carbonate (e.g. FmocVC-PAB-PNP)
An amine containing compound was dissolved in DMF and alkyl/aryl p-nitrophenyl carbonate (1.5 eq) was added, followed by addition of DIEA (2 eq) and HOBt (cat., 5%). The reaction mixture was stirred at room temperature until most of the amine was consumed. The progress of the reaction was monitored by LC/MS. The crude product was optionally purified by RP-HPLC to yield the pure carbamate.
General Procedure J—Formation of an Activated ester (e.g. NHS) from an acid
An acid was dissolved in DCM and DMF was added to aid dissolution if necessary. N-hydroxysuccinimide (1.5 eq) was added, followed by EDC·HCl (1.5 eq). The reaction mixture was stirred at room temperature for 1 h until most of the acid was consumed. The progress of the reaction was monitored by RP-HPLC. The mixture was then diluted with DCM and washed successively with citric acid (aq. 10%) and brine. The organic layer was dried and concentrated to dryness. The crude product was optionally purified by RP-HPLC or silica gel column chromatography.
Conjugation Method. Conjugation on Two Cys Residues by Forming a Cyclic Structure
Two-Step Method
Figure US12115228-20241015-C00049
One-Step Method
Figure US12115228-20241015-C00050
Triazole One Step Method
Figure US12115228-20241015-C00051
Triazine One Step Method
Figure US12115228-20241015-C00052
Experimental Description
Step 1. Drug-Linker Construct Synthesis (-L2-D)
Methods of Drug-Linker Construct Synthesis, but No Limited to:
Method 1-1: Linker and Drug Connected Via a carbamate Bond Using General Procedure G or I for Activation and carbamate Formation and General Procedure C, D, or F for Removal of Protective Groups.
Figure US12115228-20241015-C00053
Figure US12115228-20241015-C00054
Method 1-2: Linker and Drug Connected via Reductive Alkylation Reaction (General Procedure E)
Figure US12115228-20241015-C00055
Method 1-3. Active molecule containing a carboxylic acid moiety connected to an alkoxyamino linker via formation of hydoxamate (General procedure A or B), followed by removal of protective groups.
Figure US12115228-20241015-C00056
For active molecules that are hydoxamic acids, the above method still can be employed since the construct will release hydroxamic acid under enzymatic cleavage conditions. The reaction needs to start from its corresponding carboxylic acid.
Figure US12115228-20241015-C00057
Step 2. Introducing Functional Groups to L1-(L2-D)
Method 2. Introduction of o-phenylenediamine Moiety
Figure US12115228-20241015-C00058
The 3-nitro-4-amino benzoic acid was incorporated using standard amidation reaction (General procedure B). The nitro group was reduced using sodium dithionite (3 eq) in acetonitrile/water to give the desired o-phenylene diamine. MS found: 1504.8 (M+H)+.
Step 3. Introducing the final functional groups prior to conjugation
Methods of introduction of final reactive group prior to conjugation reaction, but not limited to:
Method 3-1. Formation of dibromomethyl quinoxaline
Figure US12115228-20241015-C00059
The o-phenylenediamine compound was dissolved in acetonitrile/water. Dibromomethyl diketone was added. The mixture was stirred at room temperature and purified directly by RP-HPLC to give the desired quinoxaline
Method 3-2. Formation of dichlorotriazine
Figure US12115228-20241015-C00060
To a stirred and cooled (ice bath) solution of amine compound (12 mg) in DMF (1 mL) was added a solution of cyanuric chloride (10 mg) in THF (0.5 mL) and DIEA (5 μL). The reaction mixture was stirred at room temperature for 10 mins, and then concentrated under reduced pressure. The residue was purified by RP-HPLC to give the desired dichlorotraizine (11 mg) as a white powder after lyophilization. MS found 1383.9 (M+H)+.
Method 3-3. Formation of dichloromethyltriazole
Figure US12115228-20241015-C00061
To a stirred solution of VC-PAB-MMAE (30 mg) in acetonitrile/water (2 mL, 6/4, v/v) was added a solution of trizole NHS ester (15 mg) in acetonitrile (0.5 mL) and sat aq. NaHCO3 (0.1 mL). The reaction mixture was purified by RP-HPLC to give the desired product as a white powder after lyophilization (28 mg). MS found: 1356.9 (M+H)+
Example I. Synthesis of Compound 10
Figure US12115228-20241015-C00062
To a solution of compound 1 (23.4 g, 81.53 mmol) in dry EtOH (200 ml) was added SOCl2 (100 mL) at 0° C. The mixture was stirred for overnight and the solvent was removed by evaporation in vacuo. The residue was immediately used for the next step without further purification. To a solution of compound 2 (81.53 mmol), compound 3 (50 g, 163.1 mmol) in dry DMF (150 mL) was added DEPC (15.9 g, 97.8 mmol), TEA (41 g, 0.408 mol) at 0° C. The mixture was stirred for 2 h at 0° C. Then the mixture was stirred overnight at room temperature. Solvent was removed by evaporation in vacuo. The residue was diluted with ethyl acetate-toluene (2:1, 900 ml) and washed with 1M KHSO4, water, sat. NaHCO3, and brine. The organic layer was dried and concentrated to give a residue, which was purified by column (hexanes: ethyl acetate: DCM=5:1:1) to give 38 g of compound 4.
To a solution of Boc-Val-OH (30.6 g, 0.142 mol), compound 5 (from 25 g of compound 4) in DCM (400 mL) was added BrOP (28 g, 70.87 mmol), DIEA (30 g, 0.236 mol) at 0° C. The mixture was shielded from light and stirred for 0.5 h at 0° C. Then the mixture was stirred for 48 h at room temperature. The solvent was removed by evaporation in vacuo. The residue was diluted with ethyl acetate-toluene (3:1, 900 mL) and washed with 1M KHSO4, water, sat. NaHCO3, and brine. The organic layer was dried and concentrated to give a residue, which was purified by silica gel column (hexanes: ethyl acetate: DCM=3:1:1) to give 22 g of compound 7.
To a solution of compound 7 (40 g, 66.7 mmol) in THF (600 mL) was added a mixture of LiOH (14 g, 0.33 mol) in water (300 mL) below 10° C. The mixture was stirred for 5 days at 25° C. THF was removed by evaporation. The aqueous layer was washed with Et2O (200 mL×3). The aqueous layer was acidified to pH 2 with 1N HCl at 0° C., the mixture was extracted with ethyl acetate and the organic layer was washed with water and brine. The organic layer was dried and concentrated to give a residue, which was purified by Prep-HPLC to give 14 g of compound 8.
To a solution of compound 8 (3 g) in DCM (100 mL) was added compound 9 (3 g, prepared according to General procedure J from Boc-N-Me-Val-OH using EDC and pentafluorophenol). DIEA (2.6 mL) was added, followed by HOBt (cat. 100 mg) and the reaction mixture was stirred at room temperature for 16 h. The solvents were removed under reduced pressure and the residue was purified on a silica gel column to give compound 10 as a white powder (3.1 g). MS m/z Calcd for C35H64N4O9 684.5, Found 707.6 ([M+Na]+).
Example II-1. Synthesis of Compound 13
Figure US12115228-20241015-C00063
The amino acid sulfonamide derivatives 11 were synthesized according to previously reported procedure (ARKIVOC 2004 (xii) 14-22, or WO 2007146695) using Boc protected amino acid and cyclopropyl/methyl sulfonamide, followed by removal of Boc (General procedure C)
Compound 13 was synthesized using the general procedures described above as following: DIC/HOAt mediated amide bond formation (General procedure B) between Boc-N-Me-Val-Val-Dil-Dap-OH (compound 1) and amine 11, followed by removal of Boc (General procedure C). The final compound was purified by reverse phase HPLC to give compound 13 as a white powder after lyophilization. MS m/z Calcd for C42H70N6O9S 834.5, Found 835.6 ([M+H]+).
Example II-2. Synthesis of Compounds 16
Figure US12115228-20241015-C00064
The amino acid sulfonamide derivatives 14 were synthesized according to previously reported procedure (ARKIVOC 2004 (xii) 14-22, or WO 2007146695) using Boc protected amino acid and cyclopropyl/methyl sulfonamide, followed by removal of Boc (General procedure C).
Compound 16 was synthesized using the general procedures described above as following: DIC/HOAt mediated amide bond formation (General procedure B) between Boc-N-Me-Val-Val-Dil-Dap-OH (compound 10) and amine 14, followed by removal of Boc (General procedure C). The final compound was purified by reverse phase HPLC to give compound 16 as a white powder after lyophilization. MS m/z Calcd for C41H70N6O10S 838.5, Found 839.6 ([M+H]+).
Example II-3. Synthesis of Compound 20
Figure US12115228-20241015-C00065
The phenol 16 (1 mmol) was treated with 3 eq of bis(p-nitrophenyl)carbonate to form the activated carbonate 17 (general procedure G). The crude product was used directly without further purification. 6-Aminohexanoic acid (5 eq) was dissolved in sat. aq. NaHCO3 (5 mL) and the solution was added. The reaction mixture was stirred at room temperature for 16 h. Citric acid (aq. 10%) was added to acidify the reaction (pH=4-5) and then diluted with EtOAc (150 mL). Organic layer was dried (over Na2SO4) and concentrated to give the crude product 18 which underwent the following procedures: removal of Boc (General procedure C), reductive alkylation using HCHO. The final product was purified by RP-HPLC to give compound 20 as a white powder after lyophilization. MS m/z Calcd for C48H81N7O13S 995.6, Found 996.4 ([M+H]+).
Example III. Synthesis of alkoxylamino Linkers 24, 25, 26, and 27
ID Structure
24
Figure US12115228-20241015-C00066
25
Figure US12115228-20241015-C00067
26
Figure US12115228-20241015-C00068
27
Figure US12115228-20241015-C00069
Figure US12115228-20241015-C00070
Figure US12115228-20241015-C00071
Example III-1. Synthesis of Compound 24
To a stirred solution of Fmoc-VA-PAB (21) (Bioconjugate Chem., 2002, 13, 855-859) (9 g, 15 mmol) in THF (200 mL) was added thionyl chloride (18 mmol) dropwise. After the addition was complete, the reaction mixture was stirred at room temperature for 1 h. TLC analysis (ethyl acetate/hexane, 1/1, v/v) showed the completion of the reaction. The solvents were removed under reduced pressure and the residue was washed with hexanes (100 mL) to give compound 22 as a slightly yellowish solid (8.8 g).
Compound 22 (6.2 g, 10 mmol) was dissolved in anhydrous DMF (100 mL). N-Hydroxy-phthalimide (3.2 g, 20 mmol) was added, followed by solid NaHCO3 (3.4 g, 40 mmol). The reaction mixture was stirred at room temperature for 48 h. TLC analysis showed that most of compound 61 was consumed. The reaction was then diluted with ethyl acetate (500 mL) and washed successively with sat. aq. NaHCO3 (3×200 mL) and brine (200 mL). The organic layer was dried and concentrated to give compound 23 as a tan solid, which was used directly without further purification.
The crude compound 23 from previous step was dissolved in DMF (100 mL). HOAc (6 mL) was added, followed by hydrazine hydrate (5 mL). The reaction was stirred at room temperature for 1 h. LC/MS showed the completion of the reaction. The reaction mixture was then poured into a beaker containing 1L of water under stirring. The precipitated solid was collected via filtration and washed twice with water to give compound 24 as a white solid (purity>85%, can be used directly). Pure compound 63 was obtained after RP-HPLC purification. MS m/z Calcd for C30H34N4O5 530.3, Found 531.4 ([M+H]+).
Example III-2. Synthesis of Compound 25
Compound 25 was synthesized starting from compound Fmoc-VC-PAB (Bioconjugate Chem., 2002, 13, 855-859) using the procedures described above for the synthesis of compound 25. MS m/z Calcd for C33H40N6O6 616.3, Found 617.5 ([M+H]+).
Example III-3. Synthesis of Compound 26
Compound 26 was synthesized starting from compound Fmoc-A-PAB (synthesized according to the procedure reported: Bioconjugate Chem., 2002, 13, 855-859) using the procedures described above for the synthesis of compound 26. MS m/z Calcd for C25H25N3O4 431.2, Found 432.6 ([M+H]+).
Example III-4. Synthesis of Compound 27
Compound 27 was synthesized starting from compound Fmoc-Ahx-PAB using the procedures described above for the synthesis of compound 27. MS m/z Calcd for C28H31N3O4 473.2, Found 474.3 ([M+H]+).
Example IV. Synthesis of -L1-(L2-D)
Figure US12115228-20241015-C00072
Compound 32 was synthesized using the general procedures described above as following: DIC/HOAt mediated amide bond formation (General procedure B) between Auristatin F and compound 26, followed by removal of Fmoc (General procedure D), reaction with acid 30 (General procedure A), and reduction of the nitro group to amino group.
The nitro compound 31 (32 mg) was dissolved in acetonitrile/water (6/4, v/v) and a solution of sodium dithionite (1M in water, 0.2 mL) was added. The pH of the reaction was controlled at 6-6.5 by addition of MES buffer. The mixture was stirred at room temperature for 2 h and purified directly by RP-HPLC to give the desired product as a white solid after lyophilization (18 mg). MS found 1319.0 (M+H)+.
Example V. Introduction of the Final Functional Group Prior to Conjugation Reaction
MS
found
ID Structure of —L1—(L2—D) (M + H)+
34
Figure US12115228-20241015-C00073
 		1713.0
40
Figure US12115228-20241015-C00074
 		1526.4
36
Figure US12115228-20241015-C00075
 		1383.9
39
Figure US12115228-20241015-C00076
 		1356.9
41
Figure US12115228-20241015-C00077
 		1281.2
42
Figure US12115228-20241015-C00078
 		1424.5
43
Figure US12115228-20241015-C00079
 		1249.5
44
Figure US12115228-20241015-C00080
 		1621.7
45
Figure US12115228-20241015-C00081
 		1572.7
46
Figure US12115228-20241015-C00082
 		1288.3
47
Figure US12115228-20241015-C00083
 		1510.8
48
Figure US12115228-20241015-C00084
 		1250.6
49
Figure US12115228-20241015-C00085
 		1195.4
50
Figure US12115228-20241015-C00086
 		1020.4
Example V-1. Formation of dibromomethyl quinoxaline
Figure US12115228-20241015-C00087
The o-phenylenediamine compound 32 (12 mg) was dissolved in acetonitrile/water (1 mL). Dibromomethyl diketone (10 mg) was added. The mixture was stirred at room temperature and purified directly by RP-HPLC to give the desired quinoxaline 40 as a white powder (12 mg) after lyophilization. MS found 1526.4 (M+H)+.
Compound 34, 42, 43, 44, 45, 46, and 47 were synthesized from the corresponding o-phenylenediamines using the synthetic procedure described above for the synthesis of compound 40.
Example V-2. Formation of dichlorotriazine
Figure US12115228-20241015-C00088
To a stirred and cooled solution of amine compound 35 (12 mg) in DMF (1 mL) was added a solution of cyanuric chloride (10 mg) in THF (0.5 mL) and DIEA (5 μL). The reaction mixture was stirred at room temperature for 10 mins, and then concentrated under reduced pressure. The residue was purified by RP-HPLC to give the desired dichlorotraizine 36 (11 mg) as a white powder after lyophilization. MS found 1383.9 (M+H)+.
Compound 41 was synthesized using the same procedure described above for the synthesis of compound 36. MS found 1281.2 (M+H)+.
Example V-3. Formation of dichloromethyltriazole
Figure US12115228-20241015-C00089
To a stirred solution of VC-PAB-MMAE (38, 30 mg) in acetonitrile/water (2 mL, 6/4, v/v) was added a solution of trizole NHS ester (37, 15 mg) in acetonitrile (0.5 mL) and sat. aq. NaHCO3 (0.1 mL). The reaction mixture was purified by RP-HPLC to give the desired product 39 as a white powder after lyophilization (28 mg). MS found: 1356.9 (M+H)+.
Example V-4. Synthesis of Compound 50
Figure US12115228-20241015-C00090
Fmoc-Ahx-OH (353 mg) was suspended in anhydrous DCM (5 mL) and DIC (0.5 mmol) was added. The mixture was stirred at room temperature for 1 h. Maytansinol (compound 51, 56 mg) was added to the mixture, followed by addition of 0.5 mL of pyridine. The reaction mixture was stirred at room temperature for 48 h. Solvents were removed under reduced pressure and the residue was purified by column chromatography (silica gel, EA/hexanes) to give compound 52 as a white solid (45 mg) which was treated with piperidine to remove Fmoc (General procedure D), followed by amidation reaction with 3,4-diaminobenzoic acid (1 eq) using DIC/HOBt (General procedure B, using HOBt instead of HOAt) to give compound 54 (22 mg) as a slightly yellowish powder after HPLC purification and lyophilization. Compound 54 was then converted to the desired compound 50 using the same procedure described above for the synthesis of compound 40. MS found 1020.4 (M+H)+.
Antibody-Drug Conjugation
General conjugation procedure—To a target antibody, 0.5-50 mgs/mL, in a certain buffet at pH 5.0-9.0, such as PBS, was added 0.5-100 eq of reducing agent such as TCEP and DTT. The reduction was performed at 0-40° C. for 0.5-40 hours with gentle stirring or shaking, and then the reducing agent was removed by column or ultrafiltration. To the partially reduced antibody, 0.5-50 mgs/mL, in a certain buffet at pH 5.0-9.0, such as PBS, with 0-30% of organic co-solvent such as DMA, was added 0.5-10 eq of the activated drug-linker bearing a dibromo or dichloro-functionality. The reaction was conducted at 0-40° C. for 0.5-40 hours with gentle stirring or shaking, monitored by HIC-HPLC. The resultant crude ADC product underwent necessary down-stream steps of desalt, buffet changes/formulation, and optionally, purification, using the state-of-art procedures. The final ADC product was characterized by HIC-HPLC, SEC, RP-HPLC, and optionally LC-MS. The average DAR was calculated by UV absorption and/or MS spectroscopy.
Trastuzumab and compound 34 were combined in various drug/antibody ratios under as described in the General Conjugation Procedure. Trastuzumab was incubated with 10 mM TCEP stock solution at 37° C. for 0.5 to 40 hours. After reduction, the excess of TCEP was removed by a gel filtration column. The reduced trastuzumab was then mixed with compound 34 in drug/antibody ratio range from 1 to 10 at 0 to 40° C. After overnight reaction, excess compound 34 was removed from Trastuzumab-c-34 conjugate by a gel filtration column. Purified Trastuzumab-c-34 conjugates stored at 4 to −20° C. and to be used for in vitro and in vivo tests.
The product of the conjugation reaction was analyzed using HIC-HPLC under conditions: HPLC Column: Tosoh TSKgel Butyl-NPR, 4.6 mm×3.5 cm, 2.5 mm; Buffer A: 20 mM sodium phosphate, 1.5 M Ammonium Sulfate, pH 7.0; Buffer B: 20 mM sodium phosphate, 25% v/v isopronal, pH 7.0; flow rate: 1 mL/min; Gradient: 10 min 10% Buffer B to 80% Buffer B, 4 min 100% Buffer B; 20 μL sample.
FIG. 1 shows the HIC-HPLC chromatogram product of reacting drug and antibody at a drug/antibody ratio of 5.5:1, where DAR represents the number of drugs conjugated per antibody. FIG. 2 shows the HIC-HPLC chromatogram product of reacting drug and antibody at a drug/antibody ratio of 6:1, where DAR represents the number of drugs conjugated per antibody. FIG. 3 shows the HIC-HPLC chromatogram product of reacting drug and antibody at a drug/antibody ratio of 1:6.5, where DAR represents the number of drugs conjugated per antibody. FIG. 4 shows the HIC-HPLC chromatogram product of reacting drug and antibody at a drug/antibody ratio of 1:4, where DAR represents the number of drugs conjugated per antibody. In overlay, FIG. 4 further shows HIC-HPLC chromatograms for individually isolated fractions of DAR 1, DAR 2, DAR 3, DAR 4 and DAR 5 peaks purified using FPLC. FIG. 5 shows HIC-HPLC chromatograms for each FPLC fraction, and, inset, an overlay of the peaks to demonstrate relative purity of each peak.
The purified fractions depicted in FIGS. 4 and 5 were further analyzed by reduced SDS-PAGE. The ADC samples were reduced with 20 mM DTT and loaded onto a NuPAGE 4-12% Bis-Tris Gel (Life Technology). The results are shown in FIG. 6 , where the abbreviations are as follows:
    • H-antibody heavy chain
    • L-antibody light chain
    • HH-heavy chain dimer
    • HL-heavy and light chain dimer
    • HHL-heavy-heavy and light chain trimer
    • IgG-two heavy-two light chain tetramer
      In Vitro Cytotoxicity Experiment
The antibody drug conjugates were the analyzed for cytotoxicity. The cell lines used were SK-BR-3 human breast adenocarcinoma (HER2 triple positive), HCC1954 human Ductal Carcinoma (HER2 triple positive). These cells were available from ATCC. SK-BR-3 cells were grown in McCoy's 5A medium (Caisson Labs, North Logan, UT) supplemented with 10% fetal bovine serum. HCC1954 cells were grown in RPMI-1640 medium (Caisson Labs, North Logan, UT) supplemented with 10% fetal bovine serum. SK-BR-3 cells were plated in 96-well plates at approximately 7,500 cells/well, and HCC1954 cells were plated in 96-well plates at approximately 20,000 cells/well. Antibody-drug conjugates were added in duplicates in the same day. After 72 hour incubation at 37° C., CellTiter-Glo (Promega, Madison, WI) were added and cell viability was determined as describe by the manufacture's protocol. The percent viability was determined as following:
% Viability=Average Luminescence Value of the duplicates (treated wells)/Average Luminescence Value of the untreated wells
FIG. 7 shows viability of SK-BR-3 cells for seven samples, ADC_C1 (DAR1 from FIGS. 4 and 5 ), ADC_C2 (DAR2 from FIGS. 4 and 5 ), ADC_C3 (DAR3 from FIGS. 4 and 5 ), ADC_C4 (DAR4 from FIGS. 4 and 5 ), ADC_5 (DAR5 from FIGS. 4 and 5 ), SeaGen (ADC prepared following published Seattle Genetics method), and T-DM1 (ADC prepared following published Immunogen method).
FIG. 8 shows viability of HCC1954 cells for seven samples, ADC_C1 (DAR1 from FIGS. 4 and 5 ), ADC_C2 (DAR2 from FIGS. 4 and 5 ), ADC_C3 (DAR3 from FIGS. 4 and 5 ), ADC_C4 (DAR4 from FIGS. 4 and 5 ), ADC_C5 (DAR5 from FIGS. 4 and 5 ), ADC_VC-M MAE was prepared following published Seattle Genetics method), and T-DM1 (ADC prepared following published Immunogen method).
FIG. 10 shows viability of SK-BR-3 cells for four samples, both ADC_C6 (DAR2.4) and ADC_C7 (DAR3.1) were prepared using the General Conjugation Procedure and purified by a gel filtration column. SeaGen (ADC prepared following published Seattle Genetics method), and T-DM1 (ADC prepared following published Immunogen method).
FIG. 11 shows viability of HCC1954 cells for four samples, ADC_C6 (DAR2.4 from FIG. 10 ), ADC_C7 (DAR3.1 from FIG. 10 ), ADC_VC-M MAE was prepared following published Seattle Genetics method), and T-DM1 (ADC prepared following published Immunogen method).
Antibody-Drug Conjugation
Trastuzumab and compound 42 were combined in various drug/antibody ratios under conditions where Trastuzumab was incubated with 10 mM TCEP stock solution at 37° C. for 0.5 to 40 hours. After reduction, the excess of TCEP was removed by a gel filtration column. The reduced Trastuzumab was then mixed with compound 42 in drug/antibody ratio range from 1 to 10 at 0 to 40° C. After overnight reaction, excess compound 42 was removed from Trastuzumab-c-34 conjugate by a gel filtration column. Purified Trastuzumab-c-42 conjugates stored at 4 to −20° C. and to be used for in vitro and in vivo tests.
Trastuzumab and compound 40 were combined in various drug/antibody ratios under conditions where Trastuzumab was incubated with 10 mM TCEP stock solution at 37° C. for 0.5 to 40 hours. After reduction, the excess of TCEP was removed by a gel filtration column. The reduced Trastuzumab was then mixed with compound 40 in drug/antibody ratio range from 1 to 10 at 0 to 40° C. After overnight reaction, excess compound 40 was removed from Trastuzumab-c-34 conjugate by a gel filtration column. Purified Trastuzumab-c-40 conjugates stored at 4 to −20° C. and to be used for in vitro and in vivo tests.
The product of the conjugation reaction was analyzed using reduced SDS-PAGE. The ADC samples were reduced with 20 mM DTT and loaded onto a NuPAGE 4-12% Bis-Tris Gel (Life Technology). The results are shown in FIG. 9 , where the abbreviations are as follows:
    • H-antibody heavy chain
    • L+vc-MMAE-antibody light chain-vc-MMAE
    • L-antibody light chain
    • HH-heavy chain dimer
    • HL-heavy and light chain dimer
    • HHL-heavy-heavy and light chain trimer
    • IgG-two heavy-two light chain tetramer
    • cL-Duo3: Trastuzumab-compound 42 conjugate
    • cL-Duo6: Trastuzumab-compound 40 conjugate
      In Vitro Cytotoxicity Experiment
The Trastuzumab-compound 42 and Trastuzumab-compound 42 conjugates were the analyzed for cytotoxicity. The cytotoxicity methods were performed as described above.
FIG. 12 shows viability of SK-BR-3 cells for four samples, Trastuzumab-c-Duo3 (Trastuzumab-compound 42 conjugate), Trastuzumab-c-Duo6 (Trastuzumab-compound 40 conjugate), Trastuzumab-vc-MMAE (prepared following published Seattle Genetics method), and Trastuzumab-c-MMAE (Trastuzumab-compound 34 conjugate, prepared following published Seattle Genetics method).

Claims (23)

What is claimed is:
1. An active agent-conjugate comprising Formula Ia:
Figure US12115228-20241015-C00091
or a pharmaceutically acceptable salt thereof,
wherein:
the A-component is an antibody or an antibody fragment;
the E-component is selected from the group consisting of:
Figure US12115228-20241015-C00092
L3 is an optionally substituted C1-C6 alkyl, or L3 is null, wherein when L3 is null the sulfur is directly connected to the E-component;
L4 is an optionally substituted C1-C6 alkyl, or L4 is null, wherein when L4 is null the sulfur is directly connected to the E-component;
B is a hydrophilic polymer, a biodegradable polymer, a polyamino acid, a polypeptide, a polysaccharide, a monoclonal antibody, an antibody fragment, a protein ligand, a protein scaffold, a peptide, or a nucleic acid, or B is null;
each D is independently selected, wherein each D includes an active agent;
each L2 independently comprises a —(CH2)n—, —(CH2CH2O)n—, a peptide, an oligosaccharide, or a combination thereof; wherein L2 links to E; and
n is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.
2. The active agent-conjugate of claim 1, wherein L3 is —(CH2)— and L4 is —(CH2)—.
3. The active agent-conjugate of claim 1, wherein L3 is null and L4 is null.
4. The active agent-conjugate of claim 1, wherein:
Figure US12115228-20241015-C00093
5. The active agent-conjugate of claim 1, wherein said active agent is a chemotherapy drug, a tubulin-binder, a DNA-alkylating agent, an HSP90 inhibitor, a DNA topoisomerase inhibitor, an HDAC inhibitor, a proteasome inhibitor, a peptide, an siRNA, an antisense DNA, epothilone A, epothilone B, or paclitaxel.
6. The active agent-conjugate of claim 1, wherein the A component comprises a monoclonal antibody.
7. The active agent-conjugate of claim 1, wherein the A component comprises an antibody fragment.
8. The active agent-conjugate of claim 1, wherein the A component comprises a protein ligand.
9. The active agent-conjugate of claim 1, wherein the A component comprises a protein scaffold.
10. The active agent-conjugate of claim 1, wherein the A component comprises a peptide.
11. The active agent-conjugate of claim 1, wherein the A component comprises a small molecule ligand.
12. The active agent-conjugate of claim 1, wherein the A component comprises at least one modified L-Alanine residue.
13. The active agent-conjugate of claim 1, wherein the A component comprises at least two modified L-Alanine residues.
14. The active agent-conjugate of claim 1, wherein B is null.
15. The active agent-conjugate of claim 1, wherein L2 comprises —(CH2)n—, wherein n is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.
16. The active agent-conjugate of claim 1, wherein L2 comprises —(CH2CH2O)n—, wherein n is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.
17. The active agent-conjugate of claim 1, wherein L2 comprises a peptide, an oligosaccharide, or a combination thereof.
18. The active agent-conjugate of claim 1, wherein L2 comprises (CH2)n—, —(CH2CH2O)n—, PAB, D-Val-Leu-Lys, Gly-Gly-Arg, Val-Cit-PAB, Val-Ala-PAB, Val-Lys (Ac)-PAB, Phe-Lys-PAB, Phe-Lys (Ac)-PAB, Ala-Ala-Asn-PAB, Ala-PAB, or a combination thereof.
19. The active agent-conjugate of claim 1, wherein L2 comprises a noncleavable unit.
20. The active agent-conjugate of claim 19, wherein the noncleavable unit comprises —(CH2)n—, and wherein n is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.
21. The active agent-conjugate of claim 19, wherein the noncleavable unit comprises —(CH2CH2O)n—, and wherein n is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.
22. The active agent-conjugate of claim 1, wherein L2 comprises a cleavable unit.
23. The active agent-conjugate of claim 22, wherein the cleavable unit comprises a peptide.
US17/189,135 2012-05-15 2021-03-01 Drug-conjugates, conjugation methods, and uses thereof Active 2035-02-13 US12115228B2 (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
US17/189,135 US12115228B2 (en) 2012-05-15 2021-03-01 Drug-conjugates, conjugation methods, and uses thereof

Applications Claiming Priority (8)

Application Number Priority Date Filing Date Title
US201261647300P 2012-05-15 2012-05-15
US201261648532P 2012-05-17 2012-05-17
US201261648406P 2012-05-17 2012-05-17
US201261652512P 2012-05-29 2012-05-29
PCT/US2013/041026 WO2013173391A1 (en) 2012-05-15 2013-05-14 Drug-conjugates, conjugation methods, and uses thereof
US201414401114A 2014-11-13 2014-11-13
US15/854,742 US10967071B2 (en) 2012-05-15 2017-12-26 Drug-conjugates, conjugation methods, and uses thereof
US17/189,135 US12115228B2 (en) 2012-05-15 2021-03-01 Drug-conjugates, conjugation methods, and uses thereof

Related Parent Applications (1)

Application Number Title Priority Date Filing Date
US15/854,742 Continuation US10967071B2 (en) 2012-05-15 2017-12-26 Drug-conjugates, conjugation methods, and uses thereof

Publications (2)

Publication Number Publication Date
US20210187119A1 US20210187119A1 (en) 2021-06-24
US12115228B2 true US12115228B2 (en) 2024-10-15

Family

ID=49584226

Family Applications (6)

Application Number Title Priority Date Filing Date
US14/401,318 Active US9801951B2 (en) 2012-05-15 2013-05-14 Drug-conjugates, conjugation methods, and uses thereof
US14/401,115 Active 2033-06-28 US9981046B2 (en) 2012-05-15 2013-05-14 Drug-conjugates, conjugation methods, and uses thereof
US14/401,114 Active 2033-06-04 US9884127B2 (en) 2012-05-15 2013-05-14 Drug-conjugates, conjugation methods, and uses thereof
US15/679,599 Abandoned US20180085471A1 (en) 2012-05-15 2017-08-17 Drug-conjugates, conjugation methods, and uses thereof
US15/854,742 Active 2034-06-26 US10967071B2 (en) 2012-05-15 2017-12-26 Drug-conjugates, conjugation methods, and uses thereof
US17/189,135 Active 2035-02-13 US12115228B2 (en) 2012-05-15 2021-03-01 Drug-conjugates, conjugation methods, and uses thereof

Family Applications Before (5)

Application Number Title Priority Date Filing Date
US14/401,318 Active US9801951B2 (en) 2012-05-15 2013-05-14 Drug-conjugates, conjugation methods, and uses thereof
US14/401,115 Active 2033-06-28 US9981046B2 (en) 2012-05-15 2013-05-14 Drug-conjugates, conjugation methods, and uses thereof
US14/401,114 Active 2033-06-04 US9884127B2 (en) 2012-05-15 2013-05-14 Drug-conjugates, conjugation methods, and uses thereof
US15/679,599 Abandoned US20180085471A1 (en) 2012-05-15 2017-08-17 Drug-conjugates, conjugation methods, and uses thereof
US15/854,742 Active 2034-06-26 US10967071B2 (en) 2012-05-15 2017-12-26 Drug-conjugates, conjugation methods, and uses thereof

Country Status (7)

Country Link
US (6) US9801951B2 (en)
EP (6) EP2850059A4 (en)
JP (5) JP6239597B2 (en)
CN (4) CN104379168B (en)
ES (2) ES2734259T3 (en)
HK (3) HK1208165A1 (en)
WO (3) WO2013173391A1 (en)

Families Citing this family (125)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US8288557B2 (en) 2004-07-23 2012-10-16 Endocyte, Inc. Bivalent linkers and conjugates thereof
NZ580132A (en) 2007-03-14 2012-11-30 Endocyte Inc Binding ligand linked drug delivery conjugates of tubulysins to vitamins
US9877965B2 (en) 2007-06-25 2018-01-30 Endocyte, Inc. Vitamin receptor drug delivery conjugates for treating inflammation
RU2523909C2 (en) 2007-06-25 2014-07-27 Эндосайт, Инк. Conjugates, containing hydrophilic spacers of linkers
WO2013126797A1 (en) 2012-02-24 2013-08-29 Purdue Research Foundation Cholecystokinin b receptor targeting for imaging and therapy
US20140080175A1 (en) 2012-03-29 2014-03-20 Endocyte, Inc. Processes for preparing tubulysin derivatives and conjugates thereof
US20140079636A1 (en) 2012-04-16 2014-03-20 Dinesh U. Chimmanamada Targeted therapeutics
US9801951B2 (en) 2012-05-15 2017-10-31 Concortis Biosystems, Corp. Drug-conjugates, conjugation methods, and uses thereof
AU2013331440A1 (en) 2012-10-16 2015-04-30 Endocyte, Inc. Drug delivery conjugates containing unnatural amino acids and methods for using
JP6482471B2 (en) 2012-12-21 2019-03-13 バイオアライアンス コマンディテール フェンノートシャップ Hydrophilic self-destructible linker and conjugate thereof
JP6431038B2 (en) 2013-03-15 2018-11-28 ザイムワークス インコーポレイティド Cytotoxic and antimitotic compounds and methods of use
CA2914448A1 (en) 2013-06-12 2014-12-18 Muhammad Naveed Yousaf Compounds for promoting liposomal and cellular adhesion and compositions and methods of use thereof
CA2923829A1 (en) 2013-09-10 2015-03-19 Synta Pharmaceuticals Corp. Targeted therapeutics
CA2934030A1 (en) 2013-10-15 2015-04-23 Sorrento Therapeutics Inc. Drug-conjugates with a targeting molecule and two different drugs
KR20240034882A (en) 2013-10-15 2024-03-14 씨젠 인크. Pegylated drug-linkers for improved ligand-drug conjugate pharmacokinetics
WO2015066053A2 (en) 2013-10-28 2015-05-07 Synta Pharmaceuticals Corp. Targeted therapeutics
DK3086815T3 (en) 2013-12-27 2022-05-23 Zymeworks Inc SULPHONE-CONTAINING CONNECTION SYSTEMS FOR MEDICINAL CONJUGATES
MX2016008448A (en) 2013-12-27 2017-01-09 Zymeworks Inc Var2csa-drug conjugates.
CN106132959B (en) * 2014-01-28 2020-04-17 管道制药有限责任公司 Cytotoxic tubulysin compounds for conjugation
WO2015116774A1 (en) 2014-01-29 2015-08-06 Synta Pharmaceuticals Corp. Targeted therapeutics
GB201402006D0 (en) 2014-02-06 2014-03-26 Oncomatryx Biopharma S L Antibody-drug conjugates and immunotoxins
CA2941618A1 (en) 2014-03-03 2015-09-11 Synta Pharmaceuticals Corp. Targeted therapeutics
EP3131586A4 (en) 2014-03-18 2017-10-25 Madrigal Pharmaceuticals, Inc. Targeted therapeutics
EP3149024B9 (en) * 2014-05-28 2021-10-27 Agensys, Inc. Cytotoxic dolaproine-dolaisoleuine peptide derivatives for the treatment of cancer
WO2015196167A1 (en) 2014-06-20 2015-12-23 Bioalliance C.V. Anti-folate receptor aplha (fra) antibody-drug conjugates and methods of using thereof
NZ766131A (en) 2014-07-11 2024-07-05 Genmab As Antibodies binding axl
FI3900742T3 (en) * 2014-09-11 2024-08-16 Seagen Inc Targeted delivery of tertiary amine-containing drug substances
KR102609034B1 (en) 2014-09-17 2023-12-01 자임워크스 비씨 인코포레이티드 Cytotoxic and anti-mitotic compounds, and methods of using the same
SG11201701384XA (en) 2014-10-14 2017-03-30 Polytherics Ltd Process for the conjugation of a peptide or protein with a reagent comprising a leaving group including a portion of peg
US10077287B2 (en) 2014-11-10 2018-09-18 Bristol-Myers Squibb Company Tubulysin analogs and methods of making and use
MA40725A (en) * 2014-12-08 2017-10-18 Sorrento Therapeutics Inc ANTI-C-MET-DRUG ANTIBODY CONJUGATE
US9968683B2 (en) 2014-12-17 2018-05-15 Organolinx Corp. Method for conjugating molecules
WO2016123412A1 (en) * 2015-01-28 2016-08-04 Sorrento Therapeutics, Inc. Antibody drug conjugates
WO2016127081A1 (en) * 2015-02-06 2016-08-11 Sorrento Therapeutics, Inc. Antibody drug conjugates
MX2017010102A (en) * 2015-02-15 2017-11-23 Jiangsu Hengrui Medicine Co Ligand-cytotoxicity drug conjugate, preparing method therefor, and application thereof.
CN106279352B (en) 2015-05-29 2020-05-22 上海新理念生物医药科技有限公司 Derivative of dolastatin 10 and application thereof
CN106243127B (en) 2015-06-09 2021-01-26 凯惠科技发展(上海)有限公司 Antibody drug conjugate, intermediate, preparation method, pharmaceutical composition and application
HUE049072T2 (en) 2015-07-10 2020-09-28 Genmab As Axl-specific antibody-drug conjugates for cancer treatment
CN113350518A (en) 2015-07-12 2021-09-07 杭州多禧生物科技有限公司 Conjugated bridge linkers to cell binding molecules
US9839687B2 (en) 2015-07-15 2017-12-12 Suzhou M-Conj Biotech Co., Ltd. Acetylenedicarboxyl linkers and their uses in specific conjugation of a cell-binding molecule
WO2017054080A1 (en) * 2015-09-29 2017-04-06 Zymeworks Inc. Cytotoxic and anti-mitotic compounds, and methods of using the same
US10722593B2 (en) * 2015-10-02 2020-07-28 Sirenas Llc Anti-cancer compounds and conjugates thereof
US10233212B2 (en) 2015-11-03 2019-03-19 Industrial Technology Research Institute Compounds, linker-drugs and ligand-drug conjugates
CN106729743B (en) * 2015-11-23 2021-09-21 四川科伦博泰生物医药股份有限公司 anti-ErbB 2 antibody-drug conjugate, and composition, preparation method and application thereof
US11793880B2 (en) 2015-12-04 2023-10-24 Seagen Inc. Conjugates of quaternized tubulysin compounds
EP3383420B1 (en) 2015-12-04 2022-03-23 Seagen Inc. Conjugates of quaternized tubulysin compounds
BR112018013912A2 (en) 2016-01-08 2018-12-18 Bioalliance C.V. tetravalent anti-psgl-1 antibodies and uses thereof
CN107029244B (en) * 2016-02-04 2021-04-27 浙江昭华生物医药有限公司 anti-HER 2 antibody-drug conjugate and application thereof
TW202408592A (en) 2016-03-02 2024-03-01 日商衛材R&D企管股份有限公司 Eribulin-based antibody-drug conjugates and methods of use
KR102626498B1 (en) 2016-03-25 2024-01-19 씨젠 인크. Process for the preparation of pegylated drug-linkers and intermediates thereof
WO2017172907A1 (en) * 2016-03-29 2017-10-05 Sorrento Therapeutics, Inc. Calicheamicin antibody drug conjugates linking an amidoacetyl group to a sugar moiety on calicheamicin
DK3438118T3 (en) * 2016-03-29 2023-05-30 Toray Industries Peptide derivative and use thereof
MA45328A (en) 2016-04-01 2019-02-06 Avidity Biosciences Llc NUCLEIC ACID-POLYPEPTIDE COMPOSITIONS AND USES THEREOF
CA3025931A1 (en) * 2016-05-31 2017-12-07 Sorrento Therapeutics, Inc. Antibody drug conjugates having derivatives of amatoxin as the drug
US10111966B2 (en) 2016-06-17 2018-10-30 Magenta Therapeutics, Inc. Methods for the depletion of CD117+ cells
CN108697809A (en) * 2016-08-23 2018-10-23 四川科伦博泰生物医药股份有限公司 A kind of antibody-drug conjugates and its preparation method and application
US10517958B2 (en) 2016-10-04 2019-12-31 Zymeworks Inc. Compositions and methods for the treatment of platinum-drug resistant cancer
CN109810039B (en) * 2017-11-22 2021-11-12 迈威(上海)生物科技股份有限公司 Disubstituted maleimide-based linker for antibody-drug coupling, preparation method and application thereof
CN108101825B (en) * 2016-11-25 2022-02-22 迈威(上海)生物科技股份有限公司 Disubstituted maleimide linker for antibody-drug coupling, preparation method and application thereof
EP3546448B1 (en) * 2016-11-25 2022-04-06 Mabwell (Shanghai) Bioscience Co., Ltd. Di-substituted maleic amide linker for antibody-drug conjugating and preparation method and use thereof
US10864279B2 (en) 2016-12-16 2020-12-15 Industrial Technology Research Institute Linker-drug and antibody-drug conjugate (ADC) employing the same
SG10202107429WA (en) 2017-01-06 2021-08-30 Avidity Biosciences Inc Nucleic acid-polypeptide compositions and methods of inducing exon skipping
SG10202102897PA (en) 2017-01-20 2021-04-29 Magenta Therapeutics Inc Compositions and methods for the depletion of cd137+ cells
CA3056134A1 (en) 2017-03-24 2018-09-27 Seattle Genetics, Inc. Process for the preparation of glucuronide drug-linkers and intermediates thereof
KR102655301B1 (en) * 2017-04-06 2024-04-08 항저우 디에이씨 바이오테크 씨오, 엘티디 Conjugation of a cytotoxic drug with bis-linkage
EP3617221A4 (en) 2017-04-19 2021-04-14 Sichuan Kelun-Biotech Biopharmaceutical Co., Ltd. Cytotoxin and conjugate, uses of same, and preparation method therefor
JP2020524154A (en) 2017-06-20 2020-08-13 マドリガル ファーマシューティカルズ インコーポレイテッドＭａｄｒｉｇａｌ Ｐｈａｒｍａｃｅｕｔｉｃａｌｓ，Ｉｎｃ． Targeted drug
WO2018236791A1 (en) 2017-06-20 2018-12-27 Madrigal Pharmaceuticals, Inc. Combination therapies comprising targeted therapeutics
AU2018288463A1 (en) * 2017-06-20 2020-01-30 Vivasor, Inc. CD38 antibody drug conjugate
GB201711809D0 (en) 2017-07-21 2017-09-06 Governors Of The Univ Of Alberta Antisense oligonucleotide
US10799598B2 (en) 2017-08-01 2020-10-13 Sorrento Therapeutics, Inc. Disulfide bridging conjugates
CA3072500A1 (en) 2017-08-10 2019-02-14 Sumitomo Dainippon Pharma Co., Ltd. Hemiasterlin derivatives and antibody-drug conjugates including same
EA202090670A1 (en) 2017-09-08 2020-07-28 Сиэтл Дженетикс, Инк. METHOD FOR OBTAINING TUBULISINS AND THEIR INTERMEDIATE COMPOUNDS
US11110180B2 (en) 2017-10-04 2021-09-07 Avidity Biosciences Inc. Nucleic acid-polypeptide compositions and uses thereof
US11612666B2 (en) 2017-11-30 2023-03-28 Seagen Inc. Process for the preparation of drug linker compounds
MX2020005860A (en) 2017-12-06 2020-09-09 Avidity Biosciences Inc Compositions and methods of treating muscle atrophy and myotonic dystrophy.
US11970506B2 (en) 2017-12-15 2024-04-30 Sichuan Kelun-Biotech Biopharmaceutical Co., Ltd. Bioactive molecule conjugate, preparation method and use thereof
WO2019149116A1 (en) * 2018-01-30 2019-08-08 四川科伦博泰生物医药股份有限公司 Method for preparing conjugate
MX2020008267A (en) * 2018-02-20 2020-09-21 Seattle Genetics Inc Hydrophobic auristatin f compounds and conjugates thereof.
WO2019232433A2 (en) 2018-06-01 2019-12-05 Eisai R&D Management Co., Ltd. Methods of using splicing modulators
JOP20200268A1 (en) 2018-06-01 2020-10-26 Eisai R&D Man Co Ltd Splicing modulator antibody-drug conjugates and methods of use
WO2019246445A1 (en) 2018-06-20 2019-12-26 The Research Foundation For The State University Of New York Triazamacrocycle-derived chelator compositions for coordination of imaging and therapy metal ions and methods of using same
WO2020081493A1 (en) 2018-10-16 2020-04-23 Molecular Templates, Inc. Pd-l1 binding proteins
US20220031859A1 (en) 2018-12-13 2022-02-03 Eisai R&D Management Co., Ltd. Herboxidiene antibody-drug conjugates and methods of use
IL297818B2 (en) 2018-12-21 2024-09-01 Avidity Biosciences Inc Anti-transferrin receptor antibodies and uses thereof
KR20210141918A (en) * 2018-12-21 2021-11-23 시아겐 인크. ADC with thiol multilinker
AU2020214796A1 (en) 2019-01-30 2021-07-29 Truebinding, Inc. Anti-Gal3 antibodies and uses thereof
CN109824759A (en) * 2019-03-08 2019-05-31 联宁（苏州）生物制药有限公司 A kind of Preparation Method And Their Intermediate of the drug-linker MC-MMAF for antibody drug conjugates
CA3142337A1 (en) 2019-06-06 2020-12-10 Avidity Biosciences, Inc. Una amidites and uses thereof
US11578090B2 (en) 2019-06-06 2023-02-14 Avidity Biosciences, Inc. Nucleic acid-polypeptide compositions and uses thereof
WO2021055816A1 (en) 2019-09-18 2021-03-25 Molecular Templates, Inc. Pd-l1 binding molecules comprising shiga toxin a subunit scaffolds
CN115768483A (en) 2020-01-13 2023-03-07 西纳福克斯股份有限公司 Conjugates of antibodies and immune cell adaptors
EP4090379A1 (en) 2020-01-13 2022-11-23 Synaffix B.V. Via cycloaddition bilaterally functionalized antibodies
JP2023511857A (en) 2020-01-13 2023-03-23 シンアフィックス ビー．ブイ． Antibodies bilaterally functionalized via cycloaddition
WO2021173612A1 (en) 2020-02-26 2021-09-02 Sorrento Therapeutics, Inc. Activatable antigen binding proteins with universal masking moieties
US20230129674A1 (en) * 2020-03-09 2023-04-27 Sigma-Aldrich Co. Llc Efficient preparation of dolastatin and auristatin analogs through a common intermediate
TW202144572A (en) 2020-03-19 2021-12-01 美商亞維代堤生物科學公司 Compositions and methods of treating facioscapulohumeral muscular dystrophy
TW202202173A (en) 2020-03-27 2022-01-16 美商亞維代堤生物科學公司 Compositions and methods of treating muscle dystrophy
AU2021284401A1 (en) 2020-06-05 2023-01-05 Eisai R&D Management Co., Ltd. Anti-BCMA antibody-drug conjugates and methods of use
EP4172141A1 (en) 2020-06-26 2023-05-03 Synaffix B.V. Methods for the preparation of linker payload constructs
NL2026400B1 (en) 2020-09-02 2022-05-04 Synaffix Bv Methods for the preparation of bioconjugates
JP2023540526A (en) 2020-09-04 2023-09-25 ノヴァロック バイオセラピューティクス， リミテッド Nectin-4 antibody and its use
EP4213885A1 (en) 2020-09-15 2023-07-26 Synaffix B.V. Antibody-exatecan conjugates
NL2026947B1 (en) 2020-11-20 2022-07-01 Synaffix Bv Tyrosine-based antibody conjugates
EP4267193A1 (en) 2020-12-23 2023-11-01 Ludwig-Maximilians-Universität München Improved cd30 targeting antibody drug conjugates and uses thereof
WO2022167689A1 (en) 2021-02-08 2022-08-11 Synaffix B.V. Multifunctional antibodies
CN117440832A (en) 2021-03-03 2024-01-23 索伦托药业有限公司 Antibody-drug conjugates comprising anti-BCMA antibodies
EP4308242A1 (en) 2021-03-17 2024-01-24 Molecular Templates, Inc. Pd-l1 binding proteins comprising shiga toxin a subunit scaffolds and cd8+ t cell antigens
WO2022204947A1 (en) * 2021-03-30 2022-10-06 上海复旦张江生物医药股份有限公司 Preparation method for linker drug conjugate and intermediate thereof
WO2023288252A1 (en) 2021-07-13 2023-01-19 Truebinding, Inc. Methods of preventing protein aggregation
JP2024535797A (en) 2021-09-16 2024-10-02 アビディティー バイオサイエンシーズ，インク． Compositions and methods for treating facioscapulohumeral muscular dystrophy
CA3241529A1 (en) 2021-12-23 2023-06-29 Mirecule, Inc. Compositions for delivery of polynucleotides
WO2023173026A1 (en) 2022-03-10 2023-09-14 Sorrento Therapeutics, Inc. Antibody-drug conjugates and uses thereof
US12071621B2 (en) 2022-04-05 2024-08-27 Avidity Biosciences, Inc. Anti-transferrin receptor antibody-PMO conjugates for inducing DMD exon 44 skipping
WO2024026474A1 (en) 2022-07-29 2024-02-01 Regeneron Pharmaceuticals, Inc. Compositions and methods for transferrin receptor (tfr)-mediated delivery to the brain and muscle
WO2024038065A1 (en) 2022-08-15 2024-02-22 Synaffix B.V. Anthracyclins and conjugates thereof
WO2024107765A2 (en) 2022-11-14 2024-05-23 Regeneron Pharmaceuticals, Inc. Compositions and methods for fibroblast growth factor receptor 3-mediated delivery to astrocytes
EP4389152A1 (en) 2022-12-23 2024-06-26 Synaffix B.V. Conjugates of pbd prodrugs
CN116621927B (en) * 2023-01-09 2024-03-26 联宁（苏州）生物制药有限公司 Antibody coupling intermediate with irinotecan and C-lock fixed-point coupling group, coupling method and antibody coupling drug
EP4410313A1 (en) 2023-01-31 2024-08-07 Synaffix B.V. Homogeneous antibody-conjugates with high payload loading
WO2024211236A2 (en) 2023-04-05 2024-10-10 Sorrento Therapeutics, Inc. Antibody-drug conjugates and uses thereof
WO2024211235A1 (en) 2023-04-05 2024-10-10 Sorrento Therapeutics, Inc. Antibody-drug conjugates and uses thereof
WO2024211234A1 (en) 2023-04-05 2024-10-10 Sorrento Therapeutics, Inc. Antibody-drug conjugates and uses thereof
WO2024218162A1 (en) 2023-04-17 2024-10-24 Synaffix B.V. Cleavable immune cell engagers
EP4450093A1 (en) 2023-04-17 2024-10-23 Synaffix B.V. Cleavable immune cell engagers

Citations (42)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0624377A2 (en) 1993-05-14 1994-11-17 Bristol-Myers Squibb Company Lysosomal enzyme-cleavable antitumor drug conjugates
WO1995020763A1 (en) 1994-01-31 1995-08-03 Boehringer Mannheim Corporation Piperidine analogs and conjugates of procainamide and napa
US5780588A (en) 1993-01-26 1998-07-14 Arizona Board Of Regents Elucidation and synthesis of selected pentapeptides
US6124431A (en) 1993-10-01 2000-09-26 Teikoku Hormone Mfg. Co., Ltd. Peptide derivatives
US6569834B1 (en) 1992-12-03 2003-05-27 George R. Pettit Elucidation and synthesis of antineoplastic tetrapeptide w-aminoalkyl-amides
WO2003074551A2 (en) 2002-03-01 2003-09-12 The Administrators Of The Tulane Educational Fund Conjugates of therapeutic or cytotoxic agents and biologically active peptides
WO2005081711A2 (en) 2003-11-06 2005-09-09 Seattle Genetics, Inc. Monomethylvaline compounds capable of conjugation to ligands
WO2006060533A2 (en) 2004-12-01 2006-06-08 Genentech, Inc. Conjugates of 1, 8-bis-naphthalimides with an antibody
US20060128754A1 (en) 2002-11-21 2006-06-15 Gerhard Hoefle Tubulysins, method for producing the same and tubulysin preparations
WO2007011968A2 (en) 2005-07-18 2007-01-25 Seattle Genetics, Inc. Beta-glucuronide-linker drug conjugates
US20070092940A1 (en) 2004-09-23 2007-04-26 Genentech, Inc. Cysteine engineered antibodies and conjugates
US20070134243A1 (en) 2004-12-01 2007-06-14 Gazzard Lewis J Antibody drug conjugates and methods
WO2007109567A1 (en) 2006-03-17 2007-09-27 Genentech, Inc. Anti-tat226 antibodies and immunoconjugates
WO2008019403A1 (en) 2006-08-11 2008-02-14 University Of Medicine And Dentistry Of New Jersey Dual-sensitizer-containing luminescent compounds, conjugates, and uses thereof
WO2008022309A2 (en) 2006-08-18 2008-02-21 F. Hoffmann-La Roche Ag Polyconjugates for in vivo delivery of polynucleotides
WO2008112873A2 (en) 2007-03-14 2008-09-18 Endocyte, Inc. Binding ligand linked drug delivery conjugates of tubulysins
US7531162B2 (en) 2000-10-25 2009-05-12 Mayo Foundation For Medical Education And Research Transcobalamin receptor binding conjugates useful for treating abnormal cellular proliferation
WO2010009124A2 (en) 2008-07-15 2010-01-21 Genentech, Inc. Anthracycline derivative conjugates, process for their preparation and their use as antitumor compounds
US7767205B2 (en) 2005-01-31 2010-08-03 Genentech, Inc. Anti-EphB2 antibodies and methods using same
US7829531B2 (en) 2002-07-31 2010-11-09 Seattle Genetics Inc. Drug conjugates and their use for treating cancer, an autoimmune disease or an infectious disease
JP2011515069A (en) 2008-01-31 2011-05-19 ジェネンテック， インコーポレイテッド Compositions and methods for the treatment of tumors of hematopoietic origin
US20110217321A1 (en) 2010-02-08 2011-09-08 Michael Torgov Antibody drug conjugates (adc) that bind to 161p2f10b proteins
US20110245295A1 (en) 2008-09-25 2011-10-06 Universitat Des Saarlandes Bioactive pre-tubulysins and use thereof
US20110263650A1 (en) 2007-07-20 2011-10-27 Helmholtz-Zentrum Für Infektions-Forschung Gmbh Tubulysin D Analogues
US20110268751A1 (en) 2009-01-09 2011-11-03 Seattle Genetics, Inc. Weekly Dosing Regimens for Anti-CD30 VC-PAB-MMAE Antibody Drug-Conjugates
US20110301334A1 (en) 2010-06-08 2011-12-08 Sunil Bhakta Cysteine engineered antibodies and conjugates
US8088387B2 (en) 2003-10-10 2012-01-03 Immunogen Inc. Method of targeting specific cell populations using cell-binding agent maytansinoid conjugates linked via a non-cleavable linker, said conjugates, and methods of making said conjugates
WO2012010287A1 (en) 2010-07-19 2012-01-26 Leibniz-Institut Für Pflanzenbiochemie Tubulysin analogues
CA2813056A1 (en) 2010-09-29 2012-04-05 Bayer Intellectual Property Gmbh N-carboxyalkyl auristatins and the use thereof
WO2012166559A1 (en) 2011-05-27 2012-12-06 Ambrx, Inc. Compositions containing, methods involving, and uses of non-natural amino acid linked dolastatin derivatives
WO2012166560A1 (en) 2011-05-27 2012-12-06 Ambrx, Inc. Compositions containing, methods involving, and uses of non-natural amino acid linked dolastatin derivatives
US20130029900A1 (en) 2011-06-21 2013-01-31 Immunogen, Inc. Novel maytansinoid derivatives with peptide linker and conjugates thereof
US20130101546A1 (en) 2011-06-10 2013-04-25 Mersana Therapeutics, Inc. Protein-Polymer-Drug Conjugates
WO2013085925A1 (en) 2011-12-05 2013-06-13 Igenica, Inc. Antibody-drug conjugates and related compounds, compositions, and methods
US8470984B2 (en) 2010-12-02 2013-06-25 Nerviano Medical Sciences S.R.L. Process for the preparation of morpholinyl anthracycline derivatives
WO2013173393A1 (en) 2012-05-15 2013-11-21 Concortis Biosystems, Corp Drug-conjugates, conjugation methods, and uses thereof
WO2013185117A1 (en) 2012-06-07 2013-12-12 Ambrx, Inc. Prostate-specific membrane antigen antibody drug conjugates
WO2013192360A1 (en) 2012-06-19 2013-12-27 Ambrx, Inc. Anti-cd70 antibody drug conjugates
US20140030282A1 (en) 2012-07-09 2014-01-30 Genentech, Inc. Anti-cd79b antibodies and immunoconjugates
US20150105540A1 (en) 2013-10-15 2015-04-16 Sorrento Therapeutics Inc. Drug-conjugates with a targeting molecule and two different drugs
WO2016123412A1 (en) 2015-01-28 2016-08-04 Sorrento Therapeutics, Inc. Antibody drug conjugates
WO2016127081A1 (en) 2015-02-06 2016-08-11 Sorrento Therapeutics, Inc. Antibody drug conjugates

Family Cites Families (12)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4783443A (en) 1986-03-03 1988-11-08 The University Of Chicago Amino acyl cephalosporin derivatives
EP1523493B1 (en) 2002-07-09 2013-09-04 Dömling, Alexander Novel tubulysin analogues
US7276497B2 (en) 2003-05-20 2007-10-02 Immunogen Inc. Cytotoxic agents comprising new maytansinoids
CN101087611B (en) * 2003-10-10 2019-04-23 伊缪诺金公司 With method, the conjugate and the method for preparing the conjugate of the cell binding agent maytansinoid conjugate targeting specific cells group through not cleavable connector connection
NZ566982A (en) * 2005-09-26 2011-06-30 Medarex Inc Duocarmycin drug conjugates
CL2007001630A1 (en) 2006-06-06 2008-02-08 Enanta Pharm Inc COMPOUNDS DERIVED FROM PIRROLIDINAMIDE; PHARMACEUTICAL COMPOSITION; AND USE FOR THE TREATMENT OF HEPATITIS C.
WO2008128164A1 (en) 2007-04-12 2008-10-23 The Penn State Research Foundation Accumulation field effect microelectronic device and process for the formation thereof
RU2523909C2 (en) * 2007-06-25 2014-07-27 Эндосайт, Инк. Conjugates, containing hydrophilic spacers of linkers
CN102215844A (en) * 2008-09-17 2011-10-12 恩多塞特公司 Folate receptor binding conjugates of antifolates
WO2010100430A1 (en) 2009-03-04 2010-09-10 Polytherics Limited Conjugated proteins and peptides
US8394922B2 (en) * 2009-08-03 2013-03-12 Medarex, Inc. Antiproliferative compounds, conjugates thereof, methods therefor, and uses thereof
KR20130098165A (en) * 2010-06-03 2013-09-04 제넨테크, 인크. Immuno-pet imaging of antibodies and immunoconjugates and uses therefor

Patent Citations (60)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6569834B1 (en) 1992-12-03 2003-05-27 George R. Pettit Elucidation and synthesis of antineoplastic tetrapeptide w-aminoalkyl-amides
US5780588A (en) 1993-01-26 1998-07-14 Arizona Board Of Regents Elucidation and synthesis of selected pentapeptides
EP0624377A2 (en) 1993-05-14 1994-11-17 Bristol-Myers Squibb Company Lysosomal enzyme-cleavable antitumor drug conjugates
US6124431A (en) 1993-10-01 2000-09-26 Teikoku Hormone Mfg. Co., Ltd. Peptide derivatives
WO1995020763A1 (en) 1994-01-31 1995-08-03 Boehringer Mannheim Corporation Piperidine analogs and conjugates of procainamide and napa
US7531162B2 (en) 2000-10-25 2009-05-12 Mayo Foundation For Medical Education And Research Transcobalamin receptor binding conjugates useful for treating abnormal cellular proliferation
WO2003074551A2 (en) 2002-03-01 2003-09-12 The Administrators Of The Tulane Educational Fund Conjugates of therapeutic or cytotoxic agents and biologically active peptides
CN102133407A (en) 2002-03-01 2011-07-27 图兰恩教育基金管理人 Conjugates of therapeutic or cytotoxic agents and biologically active peptides
US20110206658A1 (en) 2002-05-08 2011-08-25 Craig Crowley Compositions and methods for the treatment of tumor of hematopoietic origin
US7829531B2 (en) 2002-07-31 2010-11-09 Seattle Genetics Inc. Drug conjugates and their use for treating cancer, an autoimmune disease or an infectious disease
US20060128754A1 (en) 2002-11-21 2006-06-15 Gerhard Hoefle Tubulysins, method for producing the same and tubulysin preparations
US8088387B2 (en) 2003-10-10 2012-01-03 Immunogen Inc. Method of targeting specific cell populations using cell-binding agent maytansinoid conjugates linked via a non-cleavable linker, said conjugates, and methods of making said conjugates
US20050238649A1 (en) 2003-11-06 2005-10-27 Seattle Genetics, Inc. Monomethylvaline compounds capable of conjugation to ligands
WO2005081711A2 (en) 2003-11-06 2005-09-09 Seattle Genetics, Inc. Monomethylvaline compounds capable of conjugation to ligands
US20120148610A1 (en) 2003-11-06 2012-06-14 Seattle Genetics, Inc. Monomethylvaline compounds capable of conjugation to ligands
US7994135B2 (en) 2003-11-06 2011-08-09 Seattle Genetics, Inc. Monomethylvaline compounds capable of conjugation to ligands
US20070092940A1 (en) 2004-09-23 2007-04-26 Genentech, Inc. Cysteine engineered antibodies and conjugates
WO2006060533A2 (en) 2004-12-01 2006-06-08 Genentech, Inc. Conjugates of 1, 8-bis-naphthalimides with an antibody
US20070134243A1 (en) 2004-12-01 2007-06-14 Gazzard Lewis J Antibody drug conjugates and methods
JP2008521920A (en) 2004-12-01 2008-06-26 ジェネンテック・インコーポレーテッド Antibody drug conjugates and methods
US7767205B2 (en) 2005-01-31 2010-08-03 Genentech, Inc. Anti-EphB2 antibodies and methods using same
WO2007011968A2 (en) 2005-07-18 2007-01-25 Seattle Genetics, Inc. Beta-glucuronide-linker drug conjugates
WO2007109567A1 (en) 2006-03-17 2007-09-27 Genentech, Inc. Anti-tat226 antibodies and immunoconjugates
WO2008019403A1 (en) 2006-08-11 2008-02-14 University Of Medicine And Dentistry Of New Jersey Dual-sensitizer-containing luminescent compounds, conjugates, and uses thereof
WO2008022309A2 (en) 2006-08-18 2008-02-21 F. Hoffmann-La Roche Ag Polyconjugates for in vivo delivery of polynucleotides
WO2008112873A2 (en) 2007-03-14 2008-09-18 Endocyte, Inc. Binding ligand linked drug delivery conjugates of tubulysins
US20110263650A1 (en) 2007-07-20 2011-10-27 Helmholtz-Zentrum Für Infektions-Forschung Gmbh Tubulysin D Analogues
JP2011515069A (en) 2008-01-31 2011-05-19 ジェネンテック， インコーポレイテッド Compositions and methods for the treatment of tumors of hematopoietic origin
WO2010009124A2 (en) 2008-07-15 2010-01-21 Genentech, Inc. Anthracycline derivative conjugates, process for their preparation and their use as antitumor compounds
US20110245295A1 (en) 2008-09-25 2011-10-06 Universitat Des Saarlandes Bioactive pre-tubulysins and use thereof
US20110268751A1 (en) 2009-01-09 2011-11-03 Seattle Genetics, Inc. Weekly Dosing Regimens for Anti-CD30 VC-PAB-MMAE Antibody Drug-Conjugates
EP2376110B1 (en) 2009-01-09 2019-03-13 Seattle Genetics, Inc. Weekly dosing regimens for anti-cd30 vc-pab-mmae antibody drug-conjugates
US20110217321A1 (en) 2010-02-08 2011-09-08 Michael Torgov Antibody drug conjugates (adc) that bind to 161p2f10b proteins
US20110301334A1 (en) 2010-06-08 2011-12-08 Sunil Bhakta Cysteine engineered antibodies and conjugates
WO2012010287A1 (en) 2010-07-19 2012-01-26 Leibniz-Institut Für Pflanzenbiochemie Tubulysin analogues
CA2813056A1 (en) 2010-09-29 2012-04-05 Bayer Intellectual Property Gmbh N-carboxyalkyl auristatins and the use thereof
US8470984B2 (en) 2010-12-02 2013-06-25 Nerviano Medical Sciences S.R.L. Process for the preparation of morpholinyl anthracycline derivatives
WO2012166560A1 (en) 2011-05-27 2012-12-06 Ambrx, Inc. Compositions containing, methods involving, and uses of non-natural amino acid linked dolastatin derivatives
WO2012166559A1 (en) 2011-05-27 2012-12-06 Ambrx, Inc. Compositions containing, methods involving, and uses of non-natural amino acid linked dolastatin derivatives
US20130101546A1 (en) 2011-06-10 2013-04-25 Mersana Therapeutics, Inc. Protein-Polymer-Drug Conjugates
US20130029900A1 (en) 2011-06-21 2013-01-31 Immunogen, Inc. Novel maytansinoid derivatives with peptide linker and conjugates thereof
WO2013085925A1 (en) 2011-12-05 2013-06-13 Igenica, Inc. Antibody-drug conjugates and related compounds, compositions, and methods
JP2015500287A (en) 2011-12-05 2015-01-05 アイジェニカ・バイオセラピューティクス・インコーポレイテッドＩｇｅｎｉｃａ Ｂｉｏｔｈｅｒａｐｅｕｔｉｃｓ，Ｉｎｃ． Antibody-drug conjugates and related compounds, compositions and methods
US20130224228A1 (en) 2011-12-05 2013-08-29 Igenica, Inc. Antibody-Drug Conjugates and Related Compounds, Compositions, and Methods
US20150141646A1 (en) 2012-05-15 2015-05-21 Concortis Biosystems, Corp., a wholly owned subsidary of Sorrento Therapeutics, Inc, Drug-Conjugates, Conjugation Methods, and Uses Thereof
US9884127B2 (en) 2012-05-15 2018-02-06 Concortis Biosystems, Corp. Drug-conjugates, conjugation methods, and uses thereof
WO2013173392A1 (en) 2012-05-15 2013-11-21 Concortis Biosystems, Corp Drug-conjugates, conjugation methods, and uses thereof
WO2013173393A1 (en) 2012-05-15 2013-11-21 Concortis Biosystems, Corp Drug-conjugates, conjugation methods, and uses thereof
US20150105539A1 (en) 2012-05-15 2015-04-16 Concortis Biosystems, Corp., a wholly owned Subsidiary of Sorrento Therapeutics, Inc. Drug-Conjugates, Conjugation Methods, and Uses Thereof
WO2013173391A1 (en) 2012-05-15 2013-11-21 Concortis Biosystems, Corp Drug-conjugates, conjugation methods, and uses thereof
US20160067350A1 (en) 2012-05-15 2016-03-10 Concortis Biosystems, Corp., a wholly owned Subsidiary of Sorrento Therapeutics, Inc. Drug-Conjugates, Conjugation Methods, and Uses Thereof
US20180243438A1 (en) 2012-05-15 2018-08-30 Concortis Biosystems, Corp. Drug-conjugates, conjugation methods, and uses thereof
US9981046B2 (en) 2012-05-15 2018-05-29 Concortis Biosystems, Corp., a wholly owned Subsidiary of Sorrento Therapeutics, Inc. Drug-conjugates, conjugation methods, and uses thereof
WO2013185117A1 (en) 2012-06-07 2013-12-12 Ambrx, Inc. Prostate-specific membrane antigen antibody drug conjugates
WO2013192360A1 (en) 2012-06-19 2013-12-27 Ambrx, Inc. Anti-cd70 antibody drug conjugates
US20140030282A1 (en) 2012-07-09 2014-01-30 Genentech, Inc. Anti-cd79b antibodies and immunoconjugates
WO2015057876A1 (en) 2013-10-15 2015-04-23 Sorrento Therapeutics Inc. Drug-conjugates with a targeting molecule and two different drugs
US20150105540A1 (en) 2013-10-15 2015-04-16 Sorrento Therapeutics Inc. Drug-conjugates with a targeting molecule and two different drugs
WO2016123412A1 (en) 2015-01-28 2016-08-04 Sorrento Therapeutics, Inc. Antibody drug conjugates
WO2016127081A1 (en) 2015-02-06 2016-08-11 Sorrento Therapeutics, Inc. Antibody drug conjugates

Non-Patent Citations (31)

* Cited by examiner, † Cited by third party
Title
Antibody Structure, https://www.biology.arizona.edu/immunology/tutorials/antibody/structure.html, accessed Sep. 8, 2016.
Cella, R., et al., "Steroselective Synthesis of the Dolastatin Units by Organotriflouroborates Additions to Alpha-Amino Aldehydes", Tetrahedron Letters, 49 (2008) 16-19.
Cuellar et al (Nucleic Acids Research, 2014, vol. 1, pp. 1-15) (Year: 2014).
Dosio et al (II Farmaco, 1996, vol. 51, pp. 477-482).
Dosio et al (Journal of Pharmaceutical Sciences, 1994, vol. 83, pp. 206-211) (Year: 1994).
DubChemCompound datasheet {online compound summary) CID 56841603; Create Date: Mar. 21, 2012; https://pubchem.ncbi.nlm.nih.gov/summary/summary.cgi?cid=56841603.
Ducry, L. et al., "Antibody-Drug Conjugates: Linking Cytotoxic Payloads to Monoclonal Antibodies" Bioconjugate Chemistry, 2010, vol. 21, No. 1, pp. 5-13.
Examination Report corresponding to European Patent Application No. 18199921.0, mailed Feb. 22, 2022, 5 pages.
Goldmacher and Kovtum (Therapeutic Delivery, 2011, vol. 2, pp. 397-416) (Year: 2011).
Hollinger and Hudson, Nature Biotechnology, 2005, vol. 23, pp. 1126-1136 (Year: 2005). *
J.AM. Chem. Soc., 2010, 132, p. 1960-1965.
Kamal et al, "Remarkable DNA binding affinity and potential anticancer activity of pyrrolo[2,1-c][1,4]benzodiazepine-naphthalimide conjugates linked through piperazine side-armed alkane spacers", Bioorganic & Medicinal Chemistry, Aug. 1, 2008, vol. 16, No. 15, pp. 7218-7224. (Year: 2008). *
Kapic et al, "Synthesis of macrolones with central piperazine ring in the linker and its influence on antibacterial activity", Bioorganic & Mediicinal Chemistry: A Tetrahedron Publication for the Rapid Dissemination of Full Original Research Papers and Critical Reviews on Biomolecular Chemistry, Medicinal Chemistry and Related Disciplines, Elsevier, NL, vol. 19, No. 23, Jul. 8, 2011 (Jul. 8, 2011), pp. 7281-7298, XP028104557, ISSN: 0968-0896, DOI: 10.1016/J.BMC.2011.07.010.
Kingston, David "Tubulion Interactive Natural Products as Anticancer Agents" J Nat Prod. Mar. 2009; 72(3): 507-515.
Kovtun and Goldmacher (Cancer Letters, 2007, vol. 255, pp. 232-240) (Year: 2007).
Pettit, et al., "Specific Activities of Dolastatin 10 and Peptide Derivatives Against Crypococcus neoformans" Antimicrobial Agents and Chemotherapy, Nov. 1998, p. 2961-2965.
Pettit, G. et al., "Antineoplastic Agents. 592. Highly Effective Cancer Cell Growth Inhibitory Structural Modifications of Dolastatin 10" Journal of Natural Products, 2011, 74, 962-968.
Saavedra et al, "Piperazine as a linker for incorporating the nitric oxidereleasing diazeniumdiolate group into other biomedically relevant functional molecules", Journal of Organic Chemistry, American Chemical Society, US, vol. 64, No. 14, Jan. 1, 1999 (Jan. 1, 1999), pp. 5124-5131, XP002473170, ISSN: 0022-3263, DOI: 10.1021/J09901539.
Tannock et al., The Basic Science of Oncology, Ch. 19—Experimental Chemotherapy, p. 338 and p. 352-359, New York, 1992.
The abstract of Geroni et al (Proc. Amer. Assoc. Cancer Res., 2006, vol. 47, abstract No. 3845).
Tumey, (Next Generation Payloads for ADCs' in Innovations for the Next-Generation Antibody-Drug Conjugates, Humana Press, Cham, 2018, pp. 187-214 (Year: 2018).
U.S. Appl. No. 14/401,115, filed Nov. 13, 2014.
U.S. Appl. No. 14/401,318, filed Nov. 14, 2014.
U.S. Appl. No. 14/515,352, filed Oct. 15, 2014.
U.S. Appl. No. 15/009,775, filed Jan. 28, 2016.
U.S. Appl. No. 15/017,174, filed Feb. 5, 2016.
U.S. Appl. No. 16/886,535, filed May 28, 2020.
Uckum et al (Integrative Biology, 2013, Vo. 5, pp. 122-132) (Year: 2013).
Winkler (Therapeutic Delivery, 2013, vol. 4, pp. 791-809) (Year: 2013).
Younes, A. et al., "Brentuximab Vedotin {SGN-35) for Relapsed CD30-Positive Lymphomas" The New England Journal of Medicine, 363; 19, 2010, 1812-1821.
Zumbuehl et al., "A novel strategy for bioconjugation: synthesis and preliminary evaluation with amphotericin B", Organic & Biomolecular Chemistry, vol. 5, No. 9, Jan. 1, 2007 (Jan. 1, 2007), pp. 1339-1342.

Also Published As

Publication number Publication date
EP3590541B1 (en) 2021-08-11
US20210187119A1 (en) 2021-06-24
HK1208165A1 (en) 2016-04-01
ES2734259T3 (en) 2019-12-05
CN104640572B (en) 2018-04-27
EP3590541A1 (en) 2020-01-08
EP2849797A1 (en) 2015-03-25
US20150105539A1 (en) 2015-04-16
CN107982545A (en) 2018-05-04
CN104379168A (en) 2015-02-25
CN104662000B (en) 2018-08-17
EP3925627A1 (en) 2021-12-22
CN104640572A (en) 2015-05-20
US20180243438A1 (en) 2018-08-30
US9884127B2 (en) 2018-02-06
JP2015518833A (en) 2015-07-06
JP6239597B2 (en) 2017-11-29
US9801951B2 (en) 2017-10-31
CN107982545B (en) 2021-04-09
WO2013173391A1 (en) 2013-11-21
WO2013173392A1 (en) 2013-11-21
US9981046B2 (en) 2018-05-29
CN104379168B (en) 2017-08-15
EP2849790B1 (en) 2019-04-10
JP2018052951A (en) 2018-04-05
US20180085471A1 (en) 2018-03-29
EP2850059A1 (en) 2015-03-25
JP2018058851A (en) 2018-04-12
JP2015518831A (en) 2015-07-06
JP6239598B2 (en) 2017-11-29
EP2849790A4 (en) 2016-04-13
EP2849790A1 (en) 2015-03-25
US10967071B2 (en) 2021-04-06
US20150141646A1 (en) 2015-05-21
EP2849797B1 (en) 2019-12-18
HK1208163A1 (en) 2016-02-26
JP2015518832A (en) 2015-07-06
EP3520817A1 (en) 2019-08-07
US20160067350A1 (en) 2016-03-10
ES2887208T3 (en) 2021-12-22
EP2850059A4 (en) 2016-06-29
JP6280103B2 (en) 2018-02-14
EP2849797A4 (en) 2016-04-27
CN104662000A (en) 2015-05-27
HK1208216A1 (en) 2016-02-26
WO2013173393A1 (en) 2013-11-21

Similar Documents

Publication Publication Date Title
US12115228B2 (en) Drug-conjugates, conjugation methods, and uses thereof
US11987622B2 (en) Drug-conjugates with a targeting molecule and two different drugs
ES2804614T3 (en) Engineered Polypeptide Conjugates Using Transglutaminase
JP7257951B2 (en) Drug conjugates using self-stable linkers with improved physiochemical properties
US20050214310A1 (en) Melphalan prodrugs
US11564989B2 (en) Camptothecine antibody-drug conjugates and methods of use thereof
US20200316214A1 (en) Anti-cancer compounds and conjugates thereof
EP3665161B1 (en) New targeted cytotoxic ratjadone derivatives and conjugates thereof

Legal Events

Date Code Title Description
FEPP Fee payment procedure

Free format text: ENTITY STATUS SET TO UNDISCOUNTED (ORIGINAL EVENT CODE: BIG.); ENTITY STATUS OF PATENT OWNER: LARGE ENTITY

STPP Information on status: patent application and granting procedure in general

Free format text: APPLICATION DISPATCHED FROM PREEXAM, NOT YET DOCKETED

STPP Information on status: patent application and granting procedure in general

Free format text: DOCKETED NEW CASE - READY FOR EXAMINATION

STPP Information on status: patent application and granting procedure in general

Free format text: NON FINAL ACTION MAILED

STPP Information on status: patent application and granting procedure in general

Free format text: RESPONSE TO NON-FINAL OFFICE ACTION ENTERED AND FORWARDED TO EXAMINER

AS Assignment

Owner name: VIVASOR, INC., CALIFORNIA

Free format text: ASSIGNMENT OF ASSIGNORS INTEREST;ASSIGNOR:SORRENTO THERAPEUTICS, INC.;REEL/FRAME:067529/0019

Effective date: 20240510

STPP Information on status: patent application and granting procedure in general

Free format text: NOTICE OF ALLOWANCE MAILED -- APPLICATION RECEIVED IN OFFICE OF PUBLICATIONS

ZAAB Notice of allowance mailed

Free format text: ORIGINAL CODE: MN/=.

STPP Information on status: patent application and granting procedure in general

Free format text: AWAITING TC RESP., ISSUE FEE NOT PAID

STPP Information on status: patent application and granting procedure in general

Free format text: NOTICE OF ALLOWANCE MAILED -- APPLICATION RECEIVED IN OFFICE OF PUBLICATIONS

STPP Information on status: patent application and granting procedure in general

Free format text: NOTICE OF ALLOWANCE MAILED -- APPLICATION RECEIVED IN OFFICE OF PUBLICATIONS

STPP Information on status: patent application and granting procedure in general

Free format text: AWAITING TC RESP, ISSUE FEE PAYMENT VERIFIED

STPP Information on status: patent application and granting procedure in general

Free format text: PUBLICATIONS -- ISSUE FEE PAYMENT VERIFIED

STCF Information on status: patent grant

Free format text: PATENTED CASE